IgM-Reducing Assay Diluent is an ELISA additive specially formulated for use with problematic serum and plasma samples tested in antibody sandwich ELISA formats. This diluent can be added to standard curve/calibrator and serum/plasma sample wells to help equalize matrix complexity disparity issues.
Large differences in matrix complexity environment between standard curve and sample wells are the main cause for target analyte under-recovery problems in sandwich ELISA analysis. Proprietary additives have been included in IgM-Reducing Assay Diluent to reduce IgM-mediated conjugate bridging interference that can otherwise cause substantial background noise.
IgM-Reducing Assay Diluent contains mammalian sera proteins to reduce nonspecific interactions that might occur between the sample matrix proteins and the plate surface. It also contains a proprietary calcium chelating agent designed to inhibit interference from complement and thrombin present in serum and plasma samples. An antimicrobial agent is included in the formulation to allow for bench-top use and storage stability.
Bulk volumes and custom packaging are available upon request.
1. Once the capture antigen has been coated onto the plate and the plate has been properly blocked with one of ICT’s stabilizing blocker products, pipette 50 – 100 µL of IgM-Reducing Assay Diluent into each of the wells of the ELISA plate using a multi-channel pipettor. The addition of the Assay Diluent is always done prior to adding the standard or test samples to each well.
2. The standard and test samples should be diluted to the proper well concentrations using one of ICT’s three different Sample Diluent products to help further equalize matrix complexity parameters between standard/calibrator wells and the serum/plasma containing sample wells.
3. Because IgM-Reducing Assay Diluent contains a more diverse composition of additives, it can have some negative impact on the antigen capture step of the ELISA. This leads to reduction in ELISA sensitivity which must be compensated for by using longer antigen capture incubation times or more sensitive HRP substrate components.
• pH: 7.4 at 1X
• Contains: Mammalian Proteins
• Supplied At: 1X
• Storage: 2-8°C
Niu H, Klem T, Yang J, Qiu Y, Pan L. A biotin-drug extraction and acid dissociation (BEAD) procedure to eliminate matrix and drug interference in a protein complex anti-drug antibody (ADA) isotype specific assay. J Immunol Methods. 2017. Jul;446:30-36. Epub 2017 Apr 5. Abstract.
” – … Sulfo-tag™-conjugated anti-human/NHP IgG or IgM Ab was purchased from MSD (Meso Scale Discovery, Rockville, Maryland). IgM-reducing assay diluent was purchased from ImmunoChemistry Technologies (Bloomington, MN). … ”