Nueva llamada a la acción

Blog

 

Protein purification: the step-by-step process

Protein purification is the processes of isolating a protein of interest from its environment. In other words, from the other natural molecules surrounding the proteins in the natural niche in a host organism, or from a cell culture grown in a laboratory. Our protein purification handbook  explains that there are several available techniques and many options to consider, but the general procedure is the same. 

Read More
 

Magnetic beads vs Agarose beads: Advantages of Magnetic Agarose in Protein Purification

Most current protein purification methods use agarose beads carrying affinity functionalities such as IMAC, Glutathione, or antibodies. The choice of these functional groups depends on the protein of interest to be purified, and a large variety is available, including pre-functionalized beads that can be coupled to biomolecules (see SEPMAG® protein purification handbook chapter 4 and 5).

Read More
 

A general filtration process

Filtration is a simple technique used to separate solid particles from suspension in a liquid solution. There are many filtration methods available, but all are based on the same general principle: a heterogenous mixture is poured over a filter membrane. The filter membrane has pores of a particular size. Particles larger than the pores will be unable to pass through the membrane, while particles smaller than the pores will pass through unhindered. Additionally, all liquids will pass through. The final result of a filtration process is a collection of residue on the filtration membrane. This residue is therefore effectively separated from the rest of the mixture that passed through the membrane.

Read More
 

Upstream processing

What is upstream and downstream processing?

Introduction to upstream and downstream processing. These terms are used more in the scientific industry, for example in pharmaceutical companies. Upstream is the first half of the process and everything associated with it. Downstream is the end of the biological process. Upstream processing being the first part of the biological process, it involves the growing of bacteria in media or culturing of cell lines. Companies use bacterial or human cells to harvest products of interest. There are many products that biological companies are interested in harvesting.

Read More
 

Challenges with your Biomagnetic Separation process?

Visit Sepmag at AACR (booth 1456) and Experimental Biology (booth 1530)

SEPMAG is well known for helping IVD companies to improve, validate and scale up their biomagnetic separation processes. All this know-how on the physics behind the process also benefits researchers and industries in protein purification, cell sorting, and DNA/RNA capture.

Read More
 

Immunoprecipitation with magnetic beads

Background on Immunoprecipitation

Immuno is a prefix that means you are talking about immunity. Immunity is how the body is protected against pathogens. The immune system has a system for recognizing foreign objects, then a system for combating the presence of the foreign object. For example in humans, T-cells are a type of immune cell that recognizes antigens, structures or molecules that are foriegn. Another important immune molecule is the antibody. Antibodies are shaped like the letter Y, and the two arms of the top of the Y recognize antigens. The specific part of an antigen that is recognized is called the epitope. The antibody recognizes the epitope by its structure and sequence of amino acids. This antibody-antigen interaction serves to help the body recognize antigens. When the interaction is strong enough, it also serves as a way to neutralize antigens. Another important aspect of antibodies is that they can have highly specific interactions with an epitope, and this interaction is strong as well, also known as a high affinity interaction. These two traits, the specificity and affinity, make antibodies a great tool as well! Let’s talk more about using antibodies for a particular tool, immunoprecipitation.


Read More
 

Isolation of Cells

Techniques and investigations that require cell isolation

Cell isolation is a technique that is done in research labs and clinical settings. Cell isolation can be done in research settings to isolate a single cell to do research on it. There is a technique called patch clamp electrophysiology which measures voltage across a cell membrane. There are several ways to do this technique, either by inserting the pipette right into the membrane, or by taking a piece of the membrane off into the pipette so that molecules are still flowing through the membrane and the pipette which is connected to a device that can measure current. Another reason to isolate cells is to use them to study the effects of a drug on cell health. One grows cells in a dish in an optimized media for growth and stability. Then a drug can be introduced into the dish and one can observe how the cell physiology changes. The molecules released from the cells can also be studied or the change in the processes or proteins in the cell can also be studied with further purification or extraction techniques.


Read More
 

Magnetic properties of nanoparticles

Magnetic properties of nanoparticles are used for drug delivery, therapeutic treatment, contrast agents for MRI imaging, bioseparation, and in-vitro diagnostics.  These nanometer-sized particles are superparamagnetic, a property resulting from their tiny size—only a few nanometers—a fraction of the width of a human hair (nanoparticles are approximately 1/1,000 thinner than human hair). Superparamagnetic nanoparticles are not magnetic when located in a zero magnetic field, but they quickly become magnetized when an external magnetic field is applied. When returned to a zero magnetic field they quickly revert to a non-magnetized state. Superparamagnatism is one of the most important properties of nanoparticles used for biomagnetic separation.

Read More
 

The use of Biomagnetic Separation for improving iron oxide nanoparticles for magnetic particle imaging

Magnetic particle imaging (MPI) is a new technology that uses the signal generated by superparamagnetic tracers generated by changing magnetic fields. As it is not a natural superparamagnetic substance in the human tissues, the resultant images have no background. The tracers used in magnetic particle imaging are superparamagnetic iron oxide nanoparticles (SPIONs). The optimization of magnetic nanoparticles (MNP) plays an essential role to improve the image resolution and sensitivity of imaging techniques.

Read More
 

IVD Conference 2022

See you in Madrid! The Merck’s Annual 2022 Two-Day IVD Conference is back (and live), and we will be there...

After its virtual edition in 2021, Merck’s IVD conference will return face-to-face again on March 24-25, in Madrid, Spain. During these two days, international experts and scientists will address key aspects of critical IVD Immuno and molecular assays. Attendees will discuss current and future IVD technologies and market trends while taking advantage of networking opportunities and attending the supplier exhibition. 

Read More
 

The purpose of antibody dilution and how to do it

Antibodies are a key component to many biotechnical applications. They are most often used for immunoassays such as ELISA, cell and tissue staining, protein quantification such as western blot, and cutting edge sensor development. Verified antibodies are easily purchased from commercial vendors. These antibodies can be monoclonal or polyclonal, and can come as a lyophilized powder or as a premixed solution. All of these details must be considered when choosing which antibody to purchase because they all have an effect on the antibody concentration and dilution process. 

Read More
 

Purification Systems

Chromatography systems, or purification systems can be used to purify protein, nucleic acids, or just peptides. It comes in different sizes for different scales of purification. Research labs often do purification in smaller batches and in industry settings companies do large scale purifications. The AKTA pure is an example of one such useful technology for automating the purification process, avoiding human errors, keeping the purification at a regulated temperature such as  if you put the machine in a colder environment for less stable molecules, and having a consistent and regulated amount of pressure applied to purification columns.

Read More
 

Genomic DNA extraction

Genomic DNA extraction: an introduction

Genomic DNA is abbreviated as gDNA or called chromosomal DNA because it is packaged into chromosomes. It is the genetic code that is present in every cell and is expressed in many different combinations that lead to different cell differentiation and expression. The DNA is transcribed into RNA molecules that become proteins with many different functions. Many laboratories around the world perform genomic DNA extractions. The purpose of the genomic DNA extraction is to separate the genomic DNA from the rest of the cell contents to study it. Genomic DNA is studied by those interested in learning about specific genes and learning from genomic sequencing.


Read More
 

Nucleic Acid Labeling

Nucleic acids refer to biomolecules composed of nucleotides. A nucleotide is the name of a nucleic acid monomer which consists of a 5-carbon sugar base bound to a nitrogenous base and a phosphate group. The type of nucleotide is based on the type of nitrogenous base that is bound. For deoxyribonucleic acid (DNA) the four predominantly found bases are guanine, adenine, cytosine, and thymine. For ribonucleic acids (RNA) the four main bases are guanine, cytosine, thymine and uracil. Both DNA and RNA are part of the central dogma of molecular biology and are studied extensively.  Nucleic acids are also used for research and therapeutic purposes. In order to study and use nucleic acids, it is important to have a system of nucleic acid labeling. Nucleic acid labeling can also be used to track nucleic acids.

Read More
 

Cell separation based on cell density

The ability to isolate cells is important in both clinical and research settings. The goal of cell separation is to isolate one population of cells that are of interest. There are several reasons for performing cell separation, some examples are interest in studying a cell type or using it for therapy such as T-cell therapy. Some researchers are interested in cell separation to be used for creating hybridoma cell lines or for testing drugs in vitro and seeing the effect of the drugs on cells. There are many available techniques for cell separation. These techniques differ in specificity of cell selection, cost of equipment, time to complete, technology needed, and skill required. Cell separation based on cell density is rapid and inexpensive but is unspecific. Still, it is a fundamental technique that is commonly used in a variety of settings for general cell separation.

Read More
 

mRNA purification

mRNA purification: how mRNA vaccines work

The letters “mRNA” are heard everywhere lately. The mRNA vaccine has been widely distributed in response to COVID-19. The mRNA in the vaccine enters cells in the body where the cell machinery can translate the mRNA into the Spike surface protein of the coronavirus. The body recognizes the spike protein as an antigen and produces antibodies against it. When infected with the actual virus later on, the immune system has a base defense system, antibodies, ready to more specifically remove virus.

Read More
 

Bacterial Antigens

An antigen is defined as anything that causes an immune response in another organism. This immune response can be a simple increase of inflammatory factors, or it can be an activation of the adaptive immune system and creation of antibodies. Antibodies have two or more specific paratopes, or antigen recognition sites, that identify and combat the invading antigen. The number of antigen recognition sites is dependent on the antibody class. The word “antigen” can also refer to any protein of interest detected by a bioassay or biodetection platform. In the case of a bacterial antigen, we are referring to surface proteins, lipopolysaccharides, and peptidoglycans on the bacterial cell wall; these structures help bacteria invade other organisms by gaining access between epithelial cells. While surface structures help bacteria infect other organisms, they are also a detriment to the bacteria because they  also serve as a unique tag that antibodies and bacteriophages can recognize. Bacteriophages are viruses that attack bacteria. Both antibodies and phages are being used by scientists to develop new biodetection and biosensing platforms for rapid detection of bacterial antigens in the environment and in clinical samples.   

Read More
 

Cell centrifugation

Overview of cell biology and its importance

The main two categories of cells are prokaryotic and eukaryotic cells. Prokaryotic cells are what make up bacteria and archaea while eukaryotic cells make up organisms of the domain eukaryota. The inside of the prokaryotic cell houses it’s genetic material, DNA, in a region called the nucleoid region. There are also ribosomes within the central region of the bacteria. The next layer is the plasma membrane, a bi-lipid layer like you will see for eukaryotic cells. Unlike eukaryotic cells, prokaryotic cells will have a cell wall and a capsule surrounding the cell membrane. The eukaryotic cell is surrounded by a lipid membrane, and has membrane-bound organelles. The genetic material, DNA, is stored in the nucleus which is a membrane bound organelle. In research, many different types of cells are used. Bacterial cells are used in protein purification to grow a plasmid to express a protein of interest. You can read about this in our article protein expression and purification. Many types of Eukaryotic cells are used to do in vivo studies. Depending on your research interests, you might use muscle cells, or skin cells, or cancer cells. 

Read More
 

Magnetic particles for intracellular protein delivery

The capability of 1 μm and 2.8 μm magnetic particles to intracellularly deliver cargo proteins

In a recently published paper, researchers of the CIBER-BBN and the University Autonoma de Barcelona demonstrated that magnetic microparticles of 1 and 2.8 μm of diameter, in combination with an appropriate magnetic force, could greatly decrease the time needed to interact with and enter target cells, a clear advantage over other types of drug delivery systems.

Read More
 

Industrial uses of enzymes

Enzymes: an overview

Enzymes are proteins with the ability to catalyze chemical reactions. We have several articles you can read to learn more about proteins, their uses, and isolating them for research and clinical purposes. Check out our protein isolation article if you are thinking about how to best purify your protein of interest or read our protein assay article to learn more about working with proteins. Enzymes are particularly interesting and useful due to their catalytic activity. The molecule that goes into the enzyme for manipulation is called the substrate. You will often see this interaction called the “lock and key” interaction as the substrate has to fit just right into the pocket of the enzyme for it to work properly (enzymes are highly specific!), the way a key is very specific to the lock it goes into.

Read More
 

Affinity Chromatography and column purification of proteins and nucleic acids

Column chromatography is a method used in many areas of science to isolate a single compound from a mixture. The basic principle of column chromatography is the adsorption of target to the column by designing a column with specific affinity to the target. The target compound adsorbs to the column resin while the remaining mixture easily flow through the column and out the other end. A similar method is used to purify protein and nucleic acids, and it is generally referred to as affinity chromatography. Affinity chromatography requires a solid support (typically a magnetic bead or a resin column) on which to covalently attach a capture molecule which has affinity to the target protein or nucleic acid

Read More
 

Sonication cell lysis protocol

Cell lysis is the act of breaking the cell membrane to enable the study of specific proteins, nucleic acids, and other molecules inside of cells. When cell lysis is successful, the undamaged contents of the cell escape through the damaged cell membrane. These contents are then separated out of the mixed sample and used for further study. The methods used for separation of the lysed cell contents are dependent on the goal of the study. Careful investigation of these inner workings can reveal disease patterns, improve our understanding of normal cellular function, and elucidate biochemical pathways and therapeutic targets. Protein isolation is different from nucleic acid separation, and the reagents used vary drastically. There are a few ways to lyse the cell membrane; these include mechanical disruption, liquid homogenization, freeze/thaw cycles, manual griding, and the use of detergents. Sonication cell lysis is an example of mechanical disruption used for releasing the contents of cells.  

Read More
 

Direct and Indirect Elisa protocol

General overview of ELISA

Read More
 

Basic Guide for optimizing Chemiluminescent immunoassay (CLIA) performance and scaling-up

There are a few more considerations for optimizing the CLIA assay that in this chapter will be discussed. The following considerations are related to the performance of your assay.

Read More
 

Isolation Kit

An isolation kit helps you isolate a material of interest. When we talk about an isolation kit, we are likely talking about a kit that helps you isolate nucleic acid (RNA or DNA) or protein. These kits often contain all the buffers and hardware you need for your isolation. Let’s do a review of the types of isolation kits and how they work, then we’ll give you a general protocol to understand how the process works!

Read More
 

Chemiluminescent immunoassay (CLIA) Tracer Optimization

The tracer, the antigen or antibody labelled with a chemiluminescent tag for CLIA, is the next vital optimization step of a chemiluminescent immunoassay. As mentioned earlier, chemiluminescent labels generate light from a chemical reaction. Widely used CLIA labels  are based on luminol derivatives or acridinium esters.

Read More
 

dNTP PCR

Introduction to dNTP’s

dNTP stands for deoxynulceoside triphosphate. dNTP’s are what make up one of the four macromolecules of life, nucleic acids. A nucleoside is a molecule that consists of a ribose (sugar) bound to a nitrogenous base. On dNTP’s the ribose is actually a deoxyribose because it lacks an oxygen atom on the second carbon position. There are four dominant types of nitrogenous bases that define the type of dNTP it is, they are A,T,C,G. The triphosphate is the three phosphate groups that bind the ribose as well. Our DNA is made up of these dNTPS, A binding favorably to T and C binding favorably to G. In addition to their role in the genetics of nature, dNTP’s are also used as a tool for polymerase chain reactions (PCR). Let’s discuss how PCR works and how dTNP’s are used for it.


Read More
 

Most common Chemiluminescent immunoassay (CLIA) formats

There are several types of CLIA formats that can be used depending on the target analyte of your assay. The choice of assay format will impact four major aspects of development. The first will be the choice of magnetic bead coated with antigen/s or antibody/ies for binding the  target analyte. The tracer will then be required to match the target analyte using  a conjugated antibody/ies or antigen/s conjugatedith a CLIA label. The assay buffer will need to be optimized to improve the specificity and sensitivity for each step. Lastly there will be components such as blockers, other linking molecules or stabilizing molecules. These aspects can be optimized once an assay format is chosen. 

Read More
 

Advantages of monoclonal antibodies

Overview of antibodies

Antibodies are part of the adaptive immune response of the body. After an initial defense against a pathogen from immune cells such as neutrophils, our body launches B and T cells to create antibodies to fight a pathogen. The structure of an antibody looks like the letter Y of the latin alphabet. The central part, which goes from the stalk up to the arms of the antibody, is called the “heavy chain”. A “light chain” is attached to the upper arms of the heavy chain. The stalk is also called the “Fc” region and the arms on top are called the “Fab” region. The Fab region contains the part of the antibody that binds pathogens. This region that binds pathogens is called the paratope, and it binds an epitope on a pathogen. This interaction is specific and is based on the tertiary structure and amino acid sequence. There is one epitope per paratope.

Read More
 

Basic guide for successful magnetic bead conjugation in Chemiluminescent immunoassay (CLIA)

For a successful procedure for the magnetic bead conjugation, there are three important aspects to take in consideration when designing the assay: i) the planning of the conjugation protocol, ii) the density of the functional groups on the surface of the magnetic beads, and iii) the controlled magnetic separation of the beads.

Read More
 

Aspects of available Chemiluminescent immunoassay (CLIA) kits in the market

When developing a CLIA it is helpful to understand what is available commercially. CLIA kits are available from many different companies that formulate  reagents or components of the assay specifics for the analyte to detect and tailor made for the company analyser (platform). The solid phase can be based on superparamagnetic beads or polystyrene beads usually kept in liquid formulation. In both cases these beads require a CLIA-label reagent, discussed in more detail earlier in this ebook.

Read More
 

Streptavidin HRP

HRP stands for horseradish peroxidase, an enzyme derived from horseradish. Streptavidin is a protein derived from a species of bacteria in the genus streptomyces. Streptavidin has a high affinity for the molecule biotin. Streptavidin HRP is a streptavidin protein conjugated to HRP. HRP is used for detection/read out signals in an assay such as ELISA. Depending on which substrate you give the HRP, the enzyme will produce a different signal that you can read out with whatever hardware/technology you have in your laboratory. Let’s discuss the general protocol for using streptavidin HRP and the various substrates you can use to get an output.

Read More
 

Preliminary considerations for Chemiluminescent immunoassay (CLIA) design

Designing CLIA assays requires the consideration of different aspects, encompassing the raw materials for the reagent development & methods selection, together with the choice of the assay format. Material suppliers are a key factor for a successful design and development of an assay. An ideal supplier should be able to provide required raw materials not only at a reliable cost but also available to provide the required bulk quantities for scaling up the reagent. Moreover, suppliers should provide different lots to assess the lot to lot variability to check the impact in the assay to be developed.

Read More
 

Limitations and perspectives of Chemiluminescent immunoassay (CLIA)

Advantages of the CLIA

There are many types of assays that can be performed for detection of a molecule of interest, all with their own advantages and disadvantages. Many scientists choose to perform chemiluminescent immunoassays over the enzyme-linked immunosorbent assays (ELISA), fluorescence or radioimmunoassays. This is because the CLIA has been shown to have an improve detection at lower concentration and a wide dynamic range.

Read More
 

Spin Column

Today we are going to talk about a piece of technology found in laboratories around the world, the spin column. It is used for “solid phase extraction.” In simple terms, the spin column has a solid material that can be used to retain or bind certain molecules while letting other molecules pass through it. When referring to spin columns, this usually refers to nucleic acid purification/isolation (DNA or RNA) or proteins. The starting material can vary from blood to tissue.

Read More
 

CLIA-label protein conjugation

Once an optimal CLIA label (also called CLIA tag) is chosen, it must be conjugated to the protein which will bind to the analyte  desired to detect. Isoluminol or Acridinium ester derivatives are often used as CLIA tags. 

Read More
 

RIPA buffer recipe

General introduction to RIPA

RIPA stands for Radio immunoprecipitation assay. Let’s start out by talking about what a radio immunoprecipitation assay is and why it is used. In general, an immunoprecipitation assay uses antibodies to pull a protein of interest. The “immuno” part refers to the antibody (a molecule of the immune system) and the “precipitation” refers to a substance coming out of solution. You can read about the immunoprecipitation protocol in our other article "immunoprecipitation protocol". The radio part of this refers to using isotope labeling to track molecules. In a RIPA assay you radiolabel an antigen so you can track it when it binds an antibody, while the antibody is used to precipitate the antigen.

Read More
 

Magnetic beads in Chemiluminescent immunoassay (CLIA)

How can magnetic beads improve CLIA tests?

The combination of CLIA and magnetic beads brings together all the advantages of both parts. CLIA is known for its high sensitivity which allows the detection of analytes at very low concentrations, and thus providing an excellent limits of detection in a wide dynamic range.

Read More
 

Overview on Chemiluminescent immunoassay (CLIA)

General Introduction

Luminescence is the emission of light, and it can occur in many ways. In research and biomedical industry fluorescence and chemiluminescence are often used. Fluorescence is when light is absorbed then emitted by a substance. A photon of a higher energy state is absorbed, then a lower energy photon is emitted in another range of the electromagnetic spectrum. 

Read More
 

Protein Molarity Calculator

Quick background on proteins

Proteins are a type of macromolecule made of amino acids. Each amino acid has an amino side chain, a carboxyl group side chain and between them are the atoms unique to each amino acid, typically these unique atoms are called the “side chain” or “r group.” There are 20 amino acids that make up the chains of proteins. There are a large number of combinations that can arise from 20 amino acids, their various placement in a chain, the length of the chain, and the secondary and tertiary structure of the chain. These many combinations give rise to the diversity of proteins and their functions in nature. There are two types of secondary structure that a protein chain can take, alpha helix and beta sheet. These secondary structures further take on a tertiary structure which can happen in many ways. 

Read More
 

Monoclonal vs. Polyclonal antibodies

Easy background on antibodies

Antibodies are proteins that act as part of our adaptive immune response. In response to a pathogen, our body immediately calls on immune cells that are part of our innate immune response. The next response is called the adaptive immune response, and during this process an immune cell called a B-cell generates antibodies. The primary job of antibodies is to bind to pathogens to neutralize their ability to infect cells and to act as a tag that signals the start of more immune mechanisms.

Read More
 

ELISA standard curve

Background on ELISA

ELISA stands for enzyme-linked immunosorbant assay. To help you understand the usefulness of this technique we’ll start with a brief description of how it works. The first step is to immobilize a probe molecule to your ELISA plate, these plates are usually purchased through a vendor. A probe is a molecule that binds to a target (analyte) you are hoping to capture from a sample mixture. The probe binds to the bottom of the wells through passive adsorption to the plastic. You next add your sample and allow time for your target of interest to bind to the probe in the ELISA plate wells. Lastly, a secondary antibody is added to visualize where binding has occurred through a colorimetric or fluorescent signal. There are various versions of ELISA that modify the assay for what kind of molecule you are trying to detect in a sample and whether a primary detection antibody is available for your assay for example. You can discover which ELISA is best for you in these articles about direct and indirect ELISA, sandwich ELISA, competitive ELISA. 

Read More
 

Acetate Buffer

This blog post is going to be devoted to acetate buffer, a widely used buffer in laboratories and scientific industries. First let’s discuss acetate, a carboxylate which is the conjugate base of acetic acid (commonly known as the main component of vinegar). Acetate has a negatively charged oxygen which is why it becomes the salt, sodium acetate. In general, buffers vary in their composition because they range in their use from helping cells grow in a petri dish to stabilizing RNA for freezer conditions. Optimization of a buffer is crucial to ensure that the correct molecules are present and the pH works for the steps of your procedure in which the buffer is used.

Read More
 

Cell based assay

Why do a cell based assay?

Cell based assays are used in research and industry as an in vitro test before going to an in vivo model. For example, a company that has developed a drug has to ask several questions about how it works before even moving to any live model. The company needs to know how well their drug binds either a surface receptor of interest or many they are trying to get the drug to go into the cell. Maybe the company wants to know how well the binding of their drug to a cell triggers the desired reactions inside the cell. These types of questions refer to the potency of a drug, or the mechanism of action. Cell based assays are an ideal way to directly test how a compound or protein is interacting with a cell before trying to figure out those molecules interact with an organism as a whole.

Read More
 

Virology test

Introduction to virology and virology tests

Virologists are scientists that study viruses, which are found in astounding numbers on earth (1031!). Viruses survive only in the cells of organisms they infect, making them a parasite. Whether or not they are “living things” is a philosophical argument for a different article. One of the major reasons for studying viruses is to understand the diseases they cause in humans. Virology research has led to the development of many virology tests. It is important to diagnose as well as monitor the progression of disease of certain viruses. This means that virology tests need to be available in clinics as well as in point-of-care form to test people in areas with fewer medical resources or infrastructure. Proper diagnosis can ensure that the spread of virus can be contained and treatment can be given to those in need. A current example of a crucial virology test is SARS-CoV-2 virology tests that detect the presence of viral nucleic acid or antibodies from human samples.

Read More
 

Rna purification kit

Brief introduction to the importance of RNA

RNA (ribonucleic acids) are important macromolecules that come in many varieties and play more roles across all living organisms. Some predominant RNA are rRNA, tRNA and mRNA. tRNA is part of the process of translating proteins, rRNA is ribosomal RNA, and mRNA is messenger RNA which carries transcription information. RNA biology is an ever-growing field of research. RNA has even become a popular tool, such as the mRNA COVID-19 vaccine or CRISPR technologies. Researchers are even finding more non-coding RNA with important biological functions.

Read More
 

mRNA isolation

Introduction and inspiration of mRNA

There is a lot to learn from the mRNA (messenger RNA) that is present in cells, which makes up around 5% of the total RNA in cells. The presence of certain mRNA sequences can inform us about what proteins are most likely being translated at the moment so that biological processes can take place. mRNA has also emerged as a top vaccine for the novel coronavirus of 2019, also known as SARS-CoV-2. How can a messenger RNA be a vaccine you ask? mRNA works as a vaccine by providing our cells with the sequence to make the major protein found on SARS-CoV-2 called “spike” protein. Once our cells have made spike protein, our body will launch an immune response against it, and immune cells will make antibodies against the spike protein. To use and study mRNA, you must first perform mRNA isolation, which we will introduce in this article. 

Read More
 

RNA purification protocol

RNA (ribonucleic acid) is found in all living things. It has several functions in cells including playing a role in transcription, translation, regulation and gene expression.

Read More
 

mRNA Extraction

Overview of mRNA extraction

The “central dogma” states that DNA gets transcribed into mRNA which is then translated into a protein. The mRNA strands for messenger ribonucleic acid as it is the messenger between the directions of DNA and the creation of proteins by the ribosome. Researchers and industry laboratories extract mRNA from cells to study processes occurring in the cell. mRNA only accounts for 5% of the RNA in the cell so it is important to have a technique which will specifically purify this type of RNA. RNA is also very sensitive to Rnase contamination, which is found all over your skin as an antimicrobial. To avoid contamination it is helpful to have an efficient and simple method for mRNA extraction. A common method for mRNA extraction is the use of magnetic beads

Read More
 

Protein A vs Protein G

Protein A vs Protein G, what is the difference? They are both bacterial cell wall proteins that have primary binding sites for mammalian immunoglobulin G (IgG) antibodies, including human IgG. Protein G was first isolated from Streptococcal bacteria strains C and G. Similarly, protein A was originally found on the cell wall of the bacteria Staphylococcus aureus.  These proteins have primary binding domains for the Fc region of (IgG) antibodies, but can also recognize the Fab region of certain IgG subclasses described in more detail below.. For the bacteria this is useful because binding IgG’s at the Fc region prevents macrophages from recognizing them, which in turn prevents phagocytosis of the invading bacteria by the host immune system. For scientists this IgG binding can be used for probing in experiments such as purifications or immunoprecipitations.

Read More
 

mRNA purification kit

The purification of RNA is important for research laboratories and clinics. Throughout the COVID19 pandemic, the purification of RNA has been in the news because the rate of RNA purification defines how quickly testing for COVID19 can be done. RNA is now in the news again, mRNA specifically, as major companies have developed mRNA vaccines for COVID19. In brief, the vaccine is an injection that sends lipid coated mRNA into cells. In cells, the mRNA is translated into the spike protein of SARS-CoV-2. Companies have shown robust responses to the vaccines as the creation of the protein in cells elicits a good amount of response from the correct immune cells.

Read More
 

New MONITOR & QUALITANCE upgrade!

Version 3.2 is available (and it is FREE for all our customers!)

Read More
 

RNAi protocol

Introduction to RNA extraction kits

Read More
 

How to lyse cells

Read More
 

Working with RNA

Importance of working with RNA

Read More
 

RNA Extraction Kit

Introduction to RNA extraction kits

Read More
 

Immunoassay kits

Why do we need immunoassays kits?

There are many types of Immunoassay kits that are used in laboratories, clinical settings, and in industry. A typical laboratory bench-top kit is the ELISA kit, which stands for enzyme-linked immunosorbent assay. In a core facility or clinical setting you will find larger instrumententation for assays such as the latex turbidity immunoassay (LTIA) or chemiluminescent immunoassays (CLIA).

Read More
 

Direct Elisa and Indirect Elisa

A couple of months ago we described the sandwich elisa. Here we will discuss the other two main types of elisas—indirect and direct. Elisa is an acronym for enzyme-linked immunosorbant assay. The purpose of any elisa is to detect the presence of an antigen or antibody of interest. The indirect and direct elisa differ from the sandwich elisa because the antigen of interest is bound directly to the plate rather than a capture antibody. In either case, the key component is an enzyme-linked detection antibody. The enzyme is either colorimetric or chemiluminiscent. Chemiluminiscent enzymes are popular because they are easily read by a luminometer plate reader, making the process easy and highly quantitative.

Read More
 

Magnetic Microbead

Microbeads are composites of nanosized magnetic particles embedded in a non-magnetic matrix, the size of each bead in the range of micrometer diameter. Microbeads were originally the discovery of John Ugelstad, who was able to create uniform polystyrene spheres. Magnetic microbeads are superparamagnetic microbeads that specifically only become magnetized in the presence of a magnetic field, and this is a reversible phenomenon. These microbeads, made of materials with magnetic properties such as magnetite, have several uses in scientific research. An important use of magnetic microbeads is for separation of biological molecules from mixtures.

Read More
 

Enzymes in Industry

Enzymes play a surprisingly important role in modern industry, and are essential to the production of more commercial products than one would initially consider. Enzymes are proteins that speed up reactions and improve yield by increasing available precursors for downstream reactions. Perhaps the most obvious use for enzymes in industryisthe production of cheese, bread, and alcohol. In these traditional applications the enzymes are part of microbial machinery such as bacteria or yeast. Over time scientists have been able to isolate specific enzymes and to understand their catalytic functions well enough to incorporate them with or without their microbial hosts into a wide variety of somewhat surprising situations. For example, enzymes are used in the production of textiles, detergents, biofuels, and pharmaceutical products. Large quantities of desired enzymes are required for these applications, and they need to be available in the purest form possible. The purity of enzymes in industry is particular important for pharmaceutical applications where the products as well as the process are susceptible to review and control by regulatory associations. Batches of enzymes in industry undergo regular process validation to ensure batch-to-batch consistency.

Read More
 

Elution Buffer

An elution buffer plays an essential role in every immunoprecipitation protocol or assay that requires the release of a target antigen from a capture antibody. Elution buffers are necessary in protocols utilizing a stationary affinity column, and are also required in protocols using mobile solid supports in solution.

Read More
 

Differential Centrifugation in bio separation

The force of gravity will cause sedimentation of particles from a heterogeneous mixture; larger and denser particles sedimentfaster than the smaller and less dense particles. This phenomenon is useful for separating heterogeneous solutions into independent components, and for the isolation and enrichment of target molecules, cells, and cell organelles. Differential centrifugation accelerates the separation process by introducing centripetal forces many times greater than gravity. The precipitated particles form a pellet at the bottom of the tube during centrifugation. The rate of sedimentation is dependent on the size and density of the particles, so centrifugation can be used to isolate target particles simply by controlling centrifugal force or the rate of centrifugation. The rate of centrifugation is reported as angular velocity by revolutions per minute (rpm) or as acceleration(g). RPM is dependent on the radius of the rotor in the centrifuge.

Read More
 

Serology tests: the role of magnetic beads in the fight against COVID-19

In the fight against COVID-19, testing of patient samples has been mostly conducted using standard techniques, which has kept clinics struggling to keep up with the demand for testing. The first step in coronavirus testing that needs to be more efficient is the RNA extraction.

Download the guide: Magnetic bead coatings: Today and Tomorrow

Read More
 

The use of RNA purification magnetic kits in coronavirus testing

In a pandemic it becomes crucial to quickly design and manufacture a diagnostic device for large scale testing of human blood for viruses. Ideally, each step of the diagnosis protocol needs to scalable so that it can be done quickly. The first step, purification, needs to produce pure and clean samples for lower rates of false results. The use of RNA purification kits in coronavirus testing offers a solution to this problem. 

Read More
 

RNA magnetic purification goes large-scale

There has been a lot of discussion surrounding RNA purification for the purposes for testing people for the presence of  viruses from liquid biopsies. Using magnetic beads for the purification, many kits for individual sample preparations are required. At this time there is also potential for use of magnetic beads for large-scale purification of RNA in research towards the development of vaccines and tests. 

Read More
 

Gst tag sequence for protein isolation

Glutothione S-transferase is a 26 kDa protein that is used as an affinity tag for protein isolation in pull-down assays. The GST tag has specific affinity for the protein glutathione. This means that glutathione can be attached to columns or magnetic beads and used to isolate any protein that has been modified with the gst tag sequence. The modification of proteins with the gst tag sequence is performed in host organisms and results in fusion proteins that consist of the target protein joined by a linker to the 220 amino acids that compose the gst tag. 

Read More
 

Pharmaceutical stability testing and in-vitro diagnostics

Ensuring that pharmaceutical products reach the consumer without degradation during shipping and storage has led to the creation of stability testing guidelines. All pharmaceutical products must undergo rigorous and standardized stability tests before they are approved for sale around the world. This has not always been the case for components of In-Vitro Diagnostic (IVD) kits used in clinical and research laboratories worldwide.

Read More
 

Magnetic microbeads for cell, protein, and nucleic acid enrichment

Magnetic microbeads are used to enrich cells, proteins, and nucleic acids from complex samples using biomagnetic separation. The process requires a well-designed magnetic separation rack, and the beads need to be coated and functionalized in order to capture the desired target molecule. Magnetic microbeads are typically made of iron oxide (Fe3O4) also known as magnetite, and are 0.5 to 500 μm in diameter. The diameter of the microbeads is a function of the non-ferrous material composing the beads.(more information below). The microbeads are small enough that they are superparamagnetic, which means that they are inherently non-magnetic, but they become magnetized when placed into a magnetic field. This effect is reversible, and the magnetism disappears again after the microbeads are removed from the magnetic field.

Read More
 

Sonicator bath to resuspend aggregated magnetic beads

A sonicator bath is a tool that propagates ultrasonic waves through fluid contained within it. The ultrasonic bath is used in the laboratory to lyse cells, to degass water, and to break up clumped and aggregated magnetic beads, among many other uses. Ultrasonic cleaners are used to remove dirt and grime on objects that are hidden in difficult crevices that brushes or sprays cannot access. The most common fluid used in an ultrasonic bath or ultrasonic cleaner is distilled water. Other solvents may be added to help in cleaning processes, but in the laboratory, sonicator baths are almost always filled with distilled water. One must be careful when using solvents to ensure that they don’t have a low flash point as the ultrasonic waves will heat up the fluid in the bath. Sonicator baths work by applying ultrasonic waves to fluid. Ultrasonic waves are sound waves greater than 20 kHz; when propagated through fluid they bounce into air bubbles and cause them to burst. The shock wave released by bursting air bubbles helps to lyse cells, remove dirt from surfaces, or to break apart aggregated magnetic beads.

Read More
 

BCA protocol for protein quantification

The BCA assay is used to quantify protein concentration by using bicinchoninic acid to identify copper ions reduced by protein in a biuret reaction. The BCA protocol requires a working solution mixed with the sample; when protein is present, the reaction produces a purple color that absorbs light at 562 nm and is quantified with a spectrophotometer. The BCA assay is similar to other protein quantification assays such as Lowry or Bradford assays. However, the biuret reaction of the BCA assay occurs between the nitrogens on the peptide backbone and copper as well as nitrogens on the amino acid side chains. The fact that the peptide backbone participates in the reaction means that the BCA assay is more consistent between proteins and is less dependent upon amino acid composition. The BCA protocol is simple and quick. If the sample is heated to 37°C, then the incubation time is only 30 minutes, and the absorbance measurement takes only a few minutes. The BCA assay is an excellent method for quantifying total protein concentration after biomagnetic protein purification.

Read More
 

Multiplex Elisa immunoassay

Multiplex immunoassays enable the detection of multiple different analytes in the same sample. A well-designed multiplex assay could mean that only one vial of blood might need to be drawn from a patient instead of 8 or more vials. Therefore, one multiplex test can answer multiple questions at the same time. Immunoassays capitalize upon the specific affinity between antibody and antigen. Some immunoassays use antigen as the probe in order to detect the binding of antibody target, while other immunoassays use antibody probes in order to detect antigen targets. Most immunoassays are single-plex, meaning that they can only be used to detect one antibody-antigen pair at a time; this is generally the case with most ELISA (Enzyme Linked ImmunoSorbent Assay) tests. The idea of a multiplex ELISA is attractive, and it mostly indicates a type of multiplex immunoassay that relies upon antibody-antigen binding events. The traditional ELISA is single-plex, and requires multiple binding and washing steps as well as an enzymatic system that produces a colorimetric or chemiluminiscent label as a quantitative readout of target concentration in a sample. 

Read More
 

Sonication of cells

Sonication of cells is an essential first step to any protein purification process. Sonication is used to break apart the cell membrane, which releases all proteins into solution. Once the intracellular and transmembrane proteins are free, they can be enriched by protein purification methods. One very useful method is biomagnetic protein purification. This process uses superparamagnetic beads to isolate specific target proteins. The superparamagnetic beads are coated with proteins that specifically bind to the proteins of interest, and a magnetic separation rack attracts the beads. The remaining cellular debris is washed away and replaced by a protein isolation buffer that keeps the proteins stable until further analysis can be performed. 

Read More
 

cell in biology and drug discovery

Cellular assays form the backbone of basic research and drug discovery. A cell culture is the perfect environment in which to collect information about normal and abnormal growth, and to test novel drug compounds safely in a controlled environment. An assay in biology is carefully designed to test a single variable. Once a standard protocol for a cellular assay is established, it is highly consistent, so the chances of having confounding variables is low. This streamlines experiments and makes it easy to analyze data and draw conclusions. 

Read More
 

Magneto ELISA

A magneto elisa is a combination of magnetic bead separation and Enzyme Linked ImmunoSorbent Assay (ELISA) for analyte detection. The magnetic bead separation helps to enrich the target population from complex media such as serum or whole blood prior to quantitative detection via ELISA. This works particularly well for cell separation and detection. One example where a magneto ELISA was used, was to detect CD4+ T-cells from whole blood of HIV patients. An accurate count of CD4+ T-cells is imperative in the treatment and management of HIV and detection of AIDS development. 

Read More
 

Lateral Flow Immunoassay for qualitative and quantitative detection of protein

A lateral flow immunoassay is an easy-to-use and inexpensive paper-based device used to detect the presence of specific protein in fluid. The basic immunoassay works by taking advantage of the lock-and-key specificity of antibodies and their corresponding antigens. In the case of a lateral flow immunoassay the  capture antibodies are printed onto a paper strip and the liquid moves across it via capillary action. The presence of the target antigen is detected by a colorimetric change on the strip of paper, which also makes the lateral flow assay an example of immunochromatography. The principle component of most immunochromatography devices is usually gold nanoparticles or an enzyme-conjugated bead; the gold nanoparticles have a red hue, and enzyme conjugated beads produce a colorful product when a substrate is introduced into the system. In both instances a positive test result is visible to the naked eye. Most lateral flow immunoassays are qualitative tests, which means that a color change on the test line indicates a positive result while the lack of color indicates a negative result. There is a significant amount of research invested in the development of quantitative lateral flow immunoassays in which numerical analysis of protein concentration is possible. 

Read More
 

Isolation of CTCs from centrifuged blood and immunomagnetic separation

Circulating Tumor Cells (CTCs) are cancerous cells that dissociate from a tumor and circulate throughout the bloodstream. Therefore, the detection of CTCs in the bloodstream is an indicator of cancer progression and an early sign of metastasis. CTCs are not hematopoetic in origin, and they do not express the cell surface marker CD45. However, they do express the surface antigen EpCAM, which is commonly expressed on epithelial cells. Immunomagnetic separation methods take advantage of these surface markers to isolate CTCs from centrifuged blood samples.

Read More
 

Bioanalytical method validation

Regulation of the pharmaceutical development process is important to ensure that drug products are consistently safe and effective. There are written guidelines for pharmaceutical validation, which ensure that drug compounds are handled and tested properly. There are also separate guidelines for the bioanalytical methods used in the development and testing of new drug compounds. The goal is to standardize and improve the consistency of pharmaceutical studies and that data that are used for drug approval. Examples of analytical methods include ligand binding assays and chromatographic methods (liquid chromatography, gas chromatography, and mass spectrometry). Both the FDA (U.S. Food and Drug Administration) and EMA (European Medicines Agency) regularly update their bioanalytical method validation guidelines, but their focuses are slightly different. The FDA outlines reporting guidelines in more detail, while the EMA focuses more closely on the conduction of experiments. The validation guidelines are unified under ICH (International Council for Harmonisation of Technical requirements for pharmaceuticals for human use). 

Read More
 

How to use a sonicator with magnetic beads

Magnetic bead suspensions will eventually sediment and aggregate over time. If non-homogeneous biomagnetic separation conditions are used, studies show that the likelihood of irreversible aggregation occurring is very high. Unfortunately, when this happens, the consistency, quality and functionality of the lot are all compromised. There are several steps during the preparation of magnetic beads for diagnostic kits where irreversible aggregation can become a problem unless resuspension techniques are used that guarantee gentle disaggregation.

Read More
 

Antibody validation

Antibodies are naturally produced by the adaptive immune system in response to invading pathogens. The antibodies are made by immune cells to specifically recognize protein markers called antigens located on the outer wall or membrane of the pathogenic organism. It is this exquisite antigenic specificity that makes the adaptive immune system so remarkable in its ability to fight off a wide variety of diseases. It is also this specificity that makes the antibody-antigen interaction an attractive tool for the development of biological assays for the detection of active infection and disease.  

Read More
 

BCA protein assay

The BCA protein assay is used to quantify total protein in a biological sample. BCA stands for Bicinchoninic acid, which is the key reagent used to produce a colored product. The purple colored product is analyzed in reference to a standard curve in order to quantify protein concentration. It is important to measure protein concentration after performing a protein extraction or purification, and prior to any type of labeling procedure. The protein concentration after extraction or purification may provide information about a biochemical pathway or a disease state. All commercially available proteins are accompanied by a product information sheet that has the results of a protein quantification method. This is particular important in antibody validation. It is important to know the protein concentration prior to any labeling step so you can ensure that the stoichiometric ratio between label and protein is optimal for clean and efficient labeling. It is equally important to know how much protein you are working with when designing biosensors so that you can define limits of detection and instrument sensitivity. 

Read More
 

Process Validation Protocol

What is process validation and why do we do process validation?

Good manufacturing practice is an essential part of the production of human drugs, veterinary drugs, biological and biotechnology products, and pharmaceutical ingredients. These commercial processes are subject to regulatory oversight and must ensure that every aspect of the production process is carefully scrutinized. The purpose of process validation is to collect data and scientifically analyze the production process from conception to large scale production. An updated process validation protocol is essential to ensuring product quality and consistency. Many laws have been established to mandate process validation in order to protect consumers, especially in the case of pharmaceutical products.

Process validation in the pharmaceutical industry takes the same form as process validation in all other industries, but the stakes are higher because the product is made for human consumption. Moreover, pharmaceuticals are made to alter the natural biochemical pathways in the human body, so these chemicals must be formulated correctly and consistently every time. Additionally, the products must be stored properly and shipped under climate-controlled conditions in order to ensure efficacy once reaching the consumer.

Read More
 

Biotinylation Kit

Biotinylation kit means attaching a biotin tag to a molecule. Biotin is a natural molecule that is also known as vitamin B7. It is an important component in a healthy diet, but it is also very useful in the laboratory as a method for protein conjugation. In the laboratory, he purpose of biotinylation is to create a controlled site for biotin-streptavidin affinity binding.

What is biotin labeling and how does streptavidin bind biotin?

The biotin fits exquisitely into a biotin-binding pocket in each of the four binding sites per streptavidin molecule, and it is held in place with hydrogen bonds. Additionally, once the biotin is bound, a conformational change in the streptavidin allows a small “cap” to close over the biotin in the binding pocket. As a result, biotin and streptavidin have an extraordinary affinity for each other (Kd=10^-15). With such a low dissociation constant, once the biotin and streptavidin are bound it is unlikely that they will dissociate. This affinity is resistant to changes in temperature, pH, and salt concentration and is extremely specific. It is often thought of as a nearly covalent bond. These properties make biotinylation a useful tool for engineers who are developing new purification and detection methods. A commercially available biotinylation kit makes the process even easier.

Read More
 

Co-ip protocol: Co-immunoprecipitation

 

The difference between IP and coIP

Co-immunoprecipitation (coIP) is a protein extraction technique that specifically targets protein-protein interactions. It is slightly different from immunoprecipitation. Immunoprecipitation utilizes antibodies immobilized on a mobile support to capture target proteins. Co IP protocol takes this concept one step further by using antibodies to target not only the direct antigen that binds to the antibody, but also any protein that binds to the antigen and is pulled out with it. This makes co-ip protocol an ideal technique for studying protein complexes. The main concern when developing a co-ip protocol is to ensure that the lysis, wash, and elution buffers do not denature the proteins. Otherwise the tertiary structure of the proteins will deteriorate and the protein-protein interaction may be altered or completely lost.

Read More
 

Synthesis of fluorescent nanoparticles for bioimaging

Fluorescent nanoparticle is a general term that many people assume means any nanoscale material that produces fluorescence upon excitation with incident light. However, there is actually a thing called a conjugated polymer nanoparticle (CPN) that is very different from the fluorescent dyes (think Alexa fluorophores) that many of us are used to. These CPNs produce higher intensity fluorescence than dyes (up to 1000x brighter!), are stable and much less susceptible to quenching, and don’t contain toxic cadmium like quantum dots. These conjugated polymer fluorescent nanoparticles will likely be at the forefront of future bioimaging methods. 

Read More
 

Cell based assays

Cell based assays are used to quantify cellular function, measure how stimuli affect cells, or to localize an effect within the cell. The cells are live and intact, and require the use of fluorescent tags and chemiluminescent or colorimetric enzymes. The quantification is performed by flow cytometry or microscopy. This is very different from studies of protein or nucleic acid which require destruction of the cell and isolation of those components from cell lysate. A cell based assay is conducted entirely within live, intact cells. The goal is to understand a cellular process, localization of a molecule or drug to a cellular compartment, or to measure how cells react to a substance. Cell based assays are usually performed in tightly controlled cell lines to test for a wide range of behaviors:

Read More
 

Enlighten Biomagnetic Separation Process

What Does the Webinar Cover?

Biomagnetic separation has proven to be a quick, efficient and clean process in Life Sciences. However, most researchers and developers focus only on the magnetic beads or particles to optimize their separation process. The effectivity of the biomagnetic separation depending on the magnetic carrier is only half of the story. To have the complete picture we also need to pay attention to the role of the applied magnetic field on the play. Not understanding or controlling the parameters linked to the magnetic separator will result in failure when developing new applications, and also in industrializing lab-scale developments. The webinar will review the basic concepts of magnetic separation and help the attendees understand how advanced systems may enlight key aspects of the process. These concepts will be applied to parameterize, monitor and validate the magnetic beads behavior in controlled conditions. Afterwards, the discussion will focus on how to transfer the correctly characterized biomagnetic separation process from laboratory to production scale. Finally, the webinar will address how to use this knowledge to assure the quality of the magnetic-carriers based products.

 

New call-to-action

Read More
 

Gold nanoparticles applications

Gold nanoparticles display unique optical properties. These properties make gold nanoparticles useful tools for biotechnology and medicine. Gold nanoparticles are also called nano gold or colloidal gold due to the fact that they are less than 100nm in size and are suspended in a liquid solution. The color of the colloidal gold is dependent upon the size and shape of the gold nanoparticles comprising it. Larger particles and aggregates of particles cause the absorbance spectrum to broaden and shift towards longer wavelengths and a red color. The metallic nature of the particles makes them very useful for imaging by electron microscopy, which was one of the first applications for them. The gold nanoparticles can be functionalized with antibodies, carbohydrates, and nucleic acids. This makes them very useful for scanning electron microscopy (SEM), transmission electron microscopy (TEM), and confocal light microscopy as well as pathogen detection and other diagnostic assays. The ability of gold to absorb light via surface plasmon resonance (SPR) makes them useful tools for photothermal therapy in the treatment of cancer.

Read More
 

ELISA kit: enzyme linked immunosorbent assay

The ELISA, or enzyme linked immunosorbent assay, is the gold standard immunoassay for detection of small quantities of protein in samples as varied as serum, urine, saliva, and more. The ELISA is a labeled assay, which means that some type of label is needed to detect protein binding events. These labels are typically fluorescent, chromatic, or chemiluminiscent, and require the use of a plate reader to quantify the amount of protein in the sample. The major benefit of the ELISA is that low concentrations (often down to pg/mL) of protein are easily quantified. One disadvantage to ELISA is that many steps and reagents are required throughout the protocol. However, this can be mitigated by purchasing an ELISA kit that is pre-bound with capture antibodies and contains a detailed protocol for using all of the included buffers in a clear, easy to follow format. The use of an ELISA kit can improve diagnostic results from assay to assay because the kits are all validated between lots and come with protein standards. This means that a standard curve (detected signal vs. protein concentration) is generated during each assay and this standard curve can be checked against the expected values to ensure that the kit is still functioning as expected. The kit streamlines the process and takes the guesswork out of protocol design.

Read More
 

Choosing the appropriate DNA extraction protocol

There are three general steps to DNA extraction

  1. celllysis and deactivation of DNAases
  2. Removal of contaminating molecules: proteins, polysaccharides, salts, other nucleic acids
  3. Recovery of DNA
Read More
 

Custom antibody production

Antibodies are produced by the adaptive immune system in response to invading pathogens. The antibody has specific lock and key recognition for the offending bacteria, virus, or other molecule, which are collectively called antigens. Antibodies are proteins, which are folded polypeptides, or strands of amino acids which have antigen recognition sites that specifically recognize a binding site of its specific antigen. They are produced by B-cells of the adaptive immune system.

Read More
 

Sucrose gradient centrifugation to separate silver nanoparticles by size

As with any nanoparticle, the properties of silver nanoparticles are dependent on size. Many synthesis strategies produce nanoparticles with a wide size distribution. One way to separate these particles based on size is to use density gradient centrifugation. A sucrose gradient is an easy way to perform this separation because the sucrose gradient is simple to create using common laboratory equipment.

Read More
 

Protein Extraction Buffer

Proteins are fundamental building blocks for life. All tissues and organisms are made up of protein, and all of the work performed inside and outside of cells is mediated by protein signaling cascades. Proteins are polymers of amino acids with three or four layers of organized structure. Primary structure is defined as the linear order of amino acids. This is dictated by the genome: the code is transcribed from DNA and translated into the string of amino acids. Secondary structure is thought of as two basic forms: a beta sheet or alpha helix. The string of amino acids adopts the conformation that allows the lowest energy state. Beyond the sheets and helices, the chain can take other twists and turns to fold into a shape known as its tertiary structure. Some proteins are actually made up of two or more subunits of individually folded amino acids strands. The complexing of protein subunits to form one functional protein is called quaternary structure. All of this folding is extremely important to the character and function of each individual protein because it results in certain side chains of amino acids being located on the exterior or interior of the protein. Importantly, the folds create binding pockets where key amino acids are located to create a unique chemical landscape that allows the protein to bind to other proteins and carry out its job in a signaling cascade.

Read More
 

Gold Nanoparticle Synthesis strategies

Gold has always held a special allure for humans throughout history: wars were fought over it, alchemists strived to turn other metals into it, graves were looted for it, and love was sworn by it. Gold is so entwined in our lives that many people don’t even realize that it is fundamental to many state-of-the-art biotechnology. One of the most useful modern properties of gold is the creation of a surface plasmon resonance condition upon exposure to incident light of a resonance frequency. Gold used in this way is applied as a thin layer on a surface plasmon resonance (SPR) chip or is used as nanoparticles in Surface Enhanced Raman Scattering (SERS). These aqueous solutions of gold nanoparticles (5-50 nm in diameter) are the least recognizable form of gold because these colloidal solutions actually appear reddish in color rather than the characteristic yellow color of larger solid gold. It turns out that the work of the alchemists was not wasted. The discovery of aqua regia (noble or royal water) by an alchemist in the 8th century AD is critical to gold nanoparticle synthesis. This powerful mixture of nitric acid and hydrochloric acid is capable of dissolving solid gold to make chloroauric acid, which is the starting point for gold nanoparticle synthesis.

Read More
 

Magnetic cell separation processes in 3 steps

Cell separation is widely used in research and clinical therapy. For research purposes, sorting cells from a heterogeneous population enables the study of the different isolated types. From a therapeutic perspective, cell separation allows for the therapeutic infusion of enriched cell populations into a patient. Moreover, the latest advances in stem cell therapy, tissue engineering and regenerative medicine show the potential of cells derived from different tissues. The use of highly selective separation processes is critical to improve the quality of these cell-based treatments.

Read More
 

DC protein assay

A dc protein assay is used to quantify the amount of total protein in a sample. There are two main ways to do this, and both involve a color change as an indicator of protein presence, which means they are colorimetric assays:

Read More
 

ELISA reader

ELISA stands for Enzyme Linked Immunosorbent Assay. The immunoassay utilizes the specific lock-and-key recognition between antibodies and antigens. This recognition occurs naturally in the adaptive immune system; antibodies are created by the immune system when an antigen such as a virus or bacteria invades the body. The immune system recognizes the foreign invader and creates antibodies that specifically recognize surface proteins on the virus or bacteria. The antibodies are either attached to the surface of an immune cell or move freely through the body to tag the invader and begin a cascade of destruction and elimination. The most useful part of this process from a biotechnology and engineering perspective is the specificity of the antibody-antigen recognition.

Read More

Leave a comment