Easy-Titer™ Human IgM Assay Kit

The Thermo Scientific Easy-Titer Human IgM Assay Kit includes antibody-sensitized microspheres to measure the specific concentration of antibodies by an easy and rapid microagglutination technique using standard microplates and UV-Vis plate reader (spectrophotometer). This kit is specific for human IgM and, unlike total protein assays, can specifically measure the concentration of target antibody in samples (e.g., serum, plasma, culture supernatant) that contain other proteins. It is sensitive, requiring very small sample volumes. Antibody concentration is determined from the assay response (absorbance) by comparison to a standard curve prepared using dilutions of a known antibody sample (sold separately).

General features of Easy-Titer Antibody Assay Kits:

• Antibody-based specificity—measure concentration of target antibody in a sample, not just total protein; no need to purify antibody to assess its concentration
• Faster and easier than ELISA—three-component, homogenous assay; 10 minutes total incubation time
• No special equipment needed—uses standard vortex mixer, pipetter, 96-well microplate, plate shaker and reader (measure absorbance at 340nm or 400nm)
• Sensitive—assay range (standard curve) 8 to 500 ng/mL; use sample at 15 to 300 ng/mL for optimal results
• Reproducible—coefficient of variation less than 5%; error depends on dilution and pipetting technique
• Antibody standards sold separately—see product list for suggested products; use any antibody standard with proper target identity and known concentration (greater than 10 μg/mL)
• Kits for five popular targets—choose a kit specific for a particular species and class of immunoglobulin; no cross-reaction with other species and classes of the target antibody

Easy-Titer Assay Kits detect and measure specific target antibodies using agglutination of microspheres that are coated ('sensitized') with the specific anti-IgG or IgM polyclonal antibodies. In the appropriate aqueous buffer (supplied in kit), the monodispersed antibody-coated microspheres (> 1 μM diameter) have highest absorptivity (λ-max) to incident light having a wavelength (340nm) that is equal to approximately half their diameter. When sample is added, two or more microspheres bind to each antibody target via their coated specific polyclonal antibodies, and this agglutination into effectively larger apparent spheres results in proportional decrease in absorptivity (lower absorbance).

Typical microagglutination assays depend on a change in light-scattering and corresponding change in transmittance, to which absorbance is inversely related. Easy-Titer Assay Kits use a special dilution buffer whose refractive index eliminates the effect of light-scattering on the monodispersed microspheres for the measurement wavelength used. Because of this, the final 10- to 20-fold dilution of the sample for use in the assay must be done using the Dilution Buffer supplied in the kit.

Easy-Titer Antibody Assays are faster and easier than ELISA:
• Prepare standards (5 to 500 ng/mL) by diluting purified antibody in Kit Dilution Buffer.
• Prepare samples by diluting in Dilution Buffer to within assay range (8 to 500 ng/mL).
• Vortex vial of microsphere beads to create homogeneous suspension.
• Pipette 20 μL of bead suspension and 20 μL of each sample and standard into 96-well microplate wells.
• Incubate microplate for 5 minutes with vigorous mixing.
• Add 100 μL of Kit Blocking Reagent.
• Incubate microplate for 5 minutes with vigorous mixing.
• Measure absorbance on standard plate reader (340nm or 405nm).
• Plot standard curve and interpolate samples to determine concentration.

Related Products
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Easy-Titer™ Human IgG (H+L) Assay Kit
Easy-Titer™ Human IgG (gamma chain) Assay Kit