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There is no cure for Hepatitis B. In some cases, the infection goes away on its own (in 4 to 8 weeks for more than 9 out of 10 adults). Some (about 1 in 20 people) that contract this virus as adults become “carriers, (a chronic condition) are likely to infect other people and can do so for the rest of their lives unknowingly

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Cytokine Elisa Principle

The study of cytokines is useful in the investigation of inflammation. Cytokines are tiny proteins that convey information between cells and activate immunity in response to infection, inflammation, and other diseases. Accurate detection and measurement of cytokine concentrations are critical in characterising the nature of the infection and powerful in tracking illness progression.

Scientists are growing interested in enzyme labels and immunoassays that use enzyme-conjugated antibodies. The indirect sandwich enzyme-linked immunosorbent assay, or ELISA, is a low-cost analytical instrument used to analyse antibodies, antigens, proteins, and glycoproteins. There are numerous applications for cytokine analysis using ELISA, including those for neurological illnesses.

Helvetica Health Care (HHC) experts emphasise this significant medical application and how it aids in cytokine analysis.

What is elisa, and why is it used to detect cytokines?

The classic Sandwich ELISA test is a basic but extensively used instrument in medical laboratories throughout the world for detecting, measuring, and distinguishing a single cytokine from a mixture of biomolecules in any given sample.

ELISAs deliver specific, precise, and sensitive results, with the high-throughput rates required for cytokine protein analysis, whether the investigation is conducted in vitro or in vivo. In our whole blood assays, ELISA can typically assess cytokines produced in a culture medium, the concentration of ex vivo cytokine levels in plasma and serum, and triggered cytokine production.

Because it uses antibodies that are specific to a single cytokine or type, ELISA gives great precision. Sequential ELISA tests are used to evaluate several cytokines proteins from tiny samples.

How Does Elisas Function?

In principle, a basic sandwich ELISA analyses antigens between a layer of capture and detection antibodies. In other words, it is performed using an antibody-antigen-antibody sandwich. Below we explain the four steps of the simple indirect sandwich ELISA process:

1. Extracting analyte (specimen extracted for analysis) from a capture antibody sample.

2. Identifying biotin-labelled captured analyte with detection antibody (also specific for extracted analyte).

3. Detection amplification with streptavidin conjugated with an enzyme

4. Adding substrate and measuring the coloured signal via optical density (OD) using a microplate reader.

Scientists frequently use a microwell format (microwell plates) when conducting a traditional ELISA test.

Before we move on to a detailed explanation of the above steps, let us remind you that HHC provides a wide range of ELISA kits ready to use with break-apart wells. Our range of ELISA includes:

• The RETROTEK™ range is designed for the detection and quantitation of retroviral antigens from the retroviruses HIV-1, SIV and HTLV found in cell culture, serum, plasma or other biological fluids

• IMMUNOTEK™ kits can detect and quantify various immunoglobulins from many species, including Humans, Chicken, Cow, Goat, Rabbit, Rat and Mouse. We also provide HHV-6 IgG antibody and KSHV/HHV8 IgG antibody ELISA kits.

Describe the general elisa technique

Typically, the sample contains highly purified anti-cytokine antibodies, also known as capture antibodies, linked to an enzyme. Following plate washings, the immobilised antibodies are employed to capture the soluble cytokine proteins present in the applied samples. After washing the unbound material, biotin-conjugated anti-cytokine antibodies (detection antibodies) are utilised to identify the trapped cytokine proteins.

Following that is an enzyme-labeled avidin or streptavidin step. When a chromogenic substrate is introduced to the capture antibodies, it produces a signal that can be detected due to the substrate's colour change. The amount of coloured product produced by the bound, enzyme-linked detection reagents is simply quantified spectrophotometrically using an ELISA-plate reader at the appropriate optical density (OD). When the plate reader is connected to a computer, data storage and reanalysis are greatly facilitated.

A standard curve is introduced to a sandwich ELISA assay by serially diluting a standard cytokine protein solution with a preset concentration. Standard curves, also known as calibration curves, typically depict the standard cytokine protein concentration (usually ng or pg of cytokine/ml) vs the related mean OD value of duplicates. The concentrations of the potential cytokine-containing samples can be extrapolated from the standard curve.

The use of an ELISA computer application makes this operation easier. In general, completing a dilution series on the unknown samples helps to guarantee that the OD falls within the standard curve's linear range. The ELISA reagents used may have an impact on the curve fit analyses that researchers apply on their data, including either linear-log, log-log, or four-parameter transformations. 

Just like there are different types of cytokines, there exist other ELISAs procedures too:

· The Checkerboard method for optimising antibody concentrations

· The Dose-Response method for optimal blocking and dilution buffer for the sample matrix

· The Standard ELISA method for determining an analyte’s concentration in a given sample

· The Sequential ELISA

· The Multiplex ELISA

This fundamental approach can be improved by using sequential ELISA to test multiple analytes from a single sample aliquot. In this situation, a single sample is removed from one ELISA plate and then incubated in a different plate because the first plate would only catch the cytokine recognised by the antibody used for cytokine detection.

Multiplex ELISA is an excellent alternate method to the sequential ELISA wherein numerous capture antibodies are printed into a single microplate well, each with a different set of specificities.

Contact us at our Geneva office to inquire about the availability of our ELISA kits.