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Note: All reagents should be brought to room temperature before use.
Dilute all monoclonal antibody samples to a concentration of 1.0 ug/ml in PBS containing 1% w/v bovine serum albumin (BSA). If the concentration of the sample is entirely unknown, make dilutions based on the following estimates:
150ul of the diluted sample will be added to the development tubes.
Interpret the results at 5-10 minutes once the positive flow control bands have appeared. Within 5-10 minutes, a blue band will appear above the letters in one of the class or subclass windows as well as in the kappa window of the strip, indicating the heavy and light-chain composition of the monoclonal antibody. Lambda light chains are very rare in rat monoclonal antibodies, but if present no positive band will be seen in the kappa window. The intensity of the blue bands will increase as the sample continues to flow up the strip. The positive flow control bands on one side of the isotyping test strip should also appear, indicating that the antibody-coated micro particles are functional and have flowed up the strip. In cases where the sample is very dilute, the development time may take up to 10 minutes.
Note: For a permanent experimental record or for an easier interpretation of results when testing multiple samples, the solid red area may be cut off the bottom of the strip to prevent further band development once the positive flow control bands have appeared. A gentle stream of air can be applied to the membrane portion of the strip to assist in drying the membrane and preventing any further development. Do not wash the strip to stop the reaction.