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Mouse Immunoglobulin Isotyping ELISA Kit

Biostring > Products > Mouse Immunoglobulin Isotyping ELISA Kit
Elisa Kit

Mouse Immunoglobulin Isotyping ELISA Kit

Catalogue No BSEK-222M
Species Mouse
Size 96 T
Price POR
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SKU: BSEK-222M Categories: ,

This ELISA kit adopts the double-antibody sandwich method to identify the classes and subclasses of monoclonal antibodies in the culture supernatant of mouse lymphocyte hybridomas or specifically affinity-purified monoclonal antibodies. It can differentiate IgG1, IgG2a, IgG2b, IgG3, IgM and IgA. The microplate is pre-coated with secondary antibodies targeting the conserved epitopes of mouse immunoglobulins, which bind to mouse monoclonal antibodies in the added culture supernatant. Subsequently, HRP-labeled antibodies against mouse Ig classes and subclasses are added for specific immunoreaction. The reaction is developed with a TMB substrate system and terminated with dilute sulfuric acid. Finally, the absorbance value is measured by a microplate reader to determine the Ig class or subclass of the tested monoclonal antibody. With high resolution, this kit serves as a reliable tool for the classification and subclass identification of mouse monoclonal antibodies.

Product Composition

Name Size Name Size
Plates 96×2 Specimen dilution 15ml
Generic positive controls 1.5ml Developer A 15ml
Generic negative controls 1.5ml Color developer B 15ml
Enzyme-labeled secondary antibodies 4ml×6 Reaction Terminator 15ml
20×Cleaning solution 50mL Add sample diagram 2 pieces
Sealing film 4 pieces Instructions 1 piece

Scope of Application

Used for the identification of Ig classes and subclasses of mouse monoclonal antibodies in culture supernatant as well as related research.

Product Features

High sensitivity, high specificity, easy-to-judge results

Storage Conditions

Store at 2-8 °C away from light for 1 year if unopened. Improper storage or excessively frequent opening and use may shorten the shelf life.

1. First allow the kit to equilibrate to room temperature for approximately 30 minutes. Dilute the concentrated wash solution to working concentration with pure water at a ratio of 1 part concentrated wash solution to 19 parts pure water.

2. Take out the microplate. Allocate 6 wells for each sample, 6 wells for the positive control, and 6 wells for the negative control. Seal the unused plate strips in a self-sealing bag and place desiccant inside for preservation

3. Add 50 μL of sample diluent to each testing well first. Then add 50 μL of cell culture supernatant (or specifically affinity-purified antibody) into each of the 6 designated wells per sample. For positive control and negative control wells, add 100 μL of sample directly to each of the 6 wells without sample diluent. Cover the plate with a sealing film and incubate at 37 °C for 30 minutes

4. Discard the liquid in the wells. Wash the plate 5 times, then pat dry on lint‑free absorbent material or perform automated washing 5 times. Add 100 μL of each of the six HRP‑conjugated secondary antibodies separately into the 6 wells of each sample, as well as all positive and negative control wells. Clearly mark the corresponding wells on the plate layout diagram or the microplate. Cover with a new sealing film and incubate at 37 °C for 30 minutes.

5. Aspirate and discard the liquid in the wells. Wash the plate 5 times, then pat dry on lint‑free absorbent material or perform automated washing 5 times. Add 50 μL of Chromogen A and 50 μL of Chromogen B to each well. Cover the plate with a fresh sealing film, incubate at 37 °C in the dark for 20 minutes for color development.

6. This kit features high specificity, and results can generally be judged visually: identify the Ig class or subclass of the sample according to which well shows blue color corresponding to the conjugated secondary antibody. For accurate detection, add 50 μL of stop solution to each well to terminate the reaction, then measure absorbance with a microplate reader using dual wavelengths: 450 nm as detection wavelength and 630 nm as reference wavelength. Determine the Ig class or subclass by the secondary antibody corresponding to the high‑value well.