Human Anti-Mouse Antibody ELISA

$570.00

The Human Anti-Mouse Antibody (HAMA) ELISA Assay Kit (enzyme-linked immunoassay kit) is intended for the quantitative determination of human Anti-Mouse Antibody (HAMA) levels in serum. The Eagle Biosciences Human Anti-Mouse Antibody (HAMA) ELISA Assay Kit is for research use only and not to be used in diagnostic procedures.

SKU: HAM31-K01 Categories: , ,

Human Anti-Mouse Antibody ELISA

Human Anti-Mouse Antibody ELISA (HAMA) Developed and Manufactured in the USA

Size: 1×96 wells
Sensitivity: 2 ng/mL
Dynamic Range: 40 – 1500 ng/ml
Incubation Time: 2 hours
Sample Type: Serum, Plasma
Sample Size: 25 µL
Alternative Names: HAMA
For Research Use Only

Controls Included


Expected Normal Values

One hundred seventy normal adult sera were measured with this HAMA ELISA. One hundred sixty sera showed the OD reading very close to the zero calibrator. The 99% confidence normal cut-off is 25 ng/ml.

It is highly recommend that each laboratory establish its own normal cut off level.

Assay Principle

The Human Anti-Mouse Antibody (HAMA) ELISA Assay Kit is designed, developed and produced for the quantitative measurement of HAMA in serum and plasma samples. The assay utilizes the two-site “sandwich” technique with two selected antibodies that bind to HAMA. Assay standards, controls and patient samples are directly added to wells of a microplate that is coated with murine IgG. After the first incubation period, the HAMA binds to the murine IgG on the wall of microtiter well and unbound proteins in each microtiter well are washed away. Then a horseradish peroxidase (HRP) labeled murine IgG is added to each microtiter well and a “sandwich” of “murine IgG HAMA – murine IgG” is formed. The unbound HRP conjugated murine IgG is removed in the subsequent washing step. For the detection of this immunocomplex, the well is then incubated with a substrate solution in a timed reaction and then measured in a spectrophotometric microplate reader. The enzymatic activity of the immunocomplex bound to HAMA on the wall of the microtiter well is directly proportional to the amount of HAMA in the sample. A standard curve is generated by plotting the absorbance versus the respective HAMA concentration for each standard on point-to-point, cubical scales or 4 parameter curve fit. The concentration of HAMA in test samples is determined directly from this standard curve.


Products Related to Human Anti-Mouse Antibody ELISA

Anti-Human Antibody (HAHA) ELISA Assay Kit
Mouse Anti-Human Antibody ELISA
Anti-IgE Antibody ELISA Kit

Additional Information

Assay Background


Clinically, mouse monoclonal antibodies (IgG) and their fragments are used in vivo diagnosis procedure (radionuclides) and treatment for patients with various diseases. In patients, even a single dose injection of murine monoclonal IgG may induce immune response directed against this foreign protein (immunogen). In the circulation, the presence of human antibody against murine IgG would bind to the injected murine IgG and, therefore, diminish the efficacy of either in-vivo diagnosis or treatment. Especially, the HAMA would increase the risk of anaphylactic complications to subsequent administration of the murine IgG based therapy. The present of HAMA in patient serum or plasma specimens causes both false positive and false negative immunoassay test results depending on assay principles and monoclonal antibodies use in the assay system. This HAMA ELISA is a ready to use test kit with well-breakable microtiter plate and simple test procedures. It also provides a wide measurement range without high dose “hook” effect.

Assay Procedure


  1. Place a sufficient number of murine IgG coated microwell strips/wells in a holder to run HAMA standards, controls and unknown samples in duplicate.
  2. Add 25 µL of standards, controls and patient samples into the designated microwell.
  3. Add 100 µL of assay buffer to each well
  4. Cover the plate with one plate sealer and incubate plate at room temperature for 1 hour.
  5. Prepare HAMA Tracer antibody working solution by 1:21 fold dilution of the antibody (30196) with the tracer Antibody Diluent (30052).
  6. For each strip, it is required to mix 1 mL of the tracer antibody diluent with 50 µL of the tracer antibody in a clean test tube.
  7. Remove plate sealer. Aspirate the contents of each well. Wash each well 5 times by dispensing 350 µL of working wash solution into each well and then completely aspirating the contents. Alternatively, an automated microplate washer can be used.
  8. Add 100 µL of above diluted HAMA Tracer Antibody working solution to each of the wells.
  9. Cover the plate with the plate sealer and incubate plate at room temperature for 30 min.
  10. Remove plate sealer. Aspirate the contents of each well. Wash each well 5 times by dispensing 350 µL of working wash solution into each well and then completely aspirating the contents. Alternatively, an automated microplate washer can be used.
  11. Add 100 µL of ELISA HRP Substrate into each of the wells.
  12. Cover the plate with one plate sealer and also with aluminum foil to avoid exposure to light.
  13. Incubate plate at room temperature for 20 min.
  14. Remove the aluminum foil and plate sealer. Add 100 µL of ELISA Stop Solution into each of the wells. Mix gently.
  15. Read the absorbance at 450 nm within 10 minutes in a microplate reader

Typical Standard Curve


Human Anti-Mouse Antibody ELISA Standard Curve

Documents

Product Documents


 

Please note: All documents above are for reference use only and should not be used in place of the documents included with this physical product. If digital copies are needed, please contact us.

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