Immunoassay Optimization Antibody and Sample Diluent
1. What is special about LowCross-Buffer®?
LowCross-Buffer® is a sample buffer for immunoassays that minimizes unwanted interference, such as non-specific binding, cross-reactivities and matrix effects. The high affinity, specific binding of the antibody to the analyte is retained, while undesired binding of the antibody is prevented.
2. How does LowCross-Buffer® reduce matrix effects?
Matrix effects usually result from unwanted low or medium affinity binding of the matrix components, such as serum proteins from human serum specimen, to the analyte or antibody. If the analyte is masked by proteins or other components of the specimen, the antibody cannot bind to the analyte. Antibodies can be masked by matrix components as well. LowCross-Buffer® reduces this masking of antibodies.
3. How is LowCross-Buffer® used?
LowCross-Buffer® is a dilution buffer for specimens, such as serum or plasma, and/or for detection antibodies. Its use depends on the interference. If the non-specific binding or cross-reactivity derives from the detection antibody, we recommend diluting the detection antibody in LowCross-Buffer®.
4. Does LowCross-Buffer® need to be diluted?
No. LowCross-Buffer® is ready-to-use. Sample or detection antibodies are diluted directly with the buffer. In competitive assays and some applications of immunohistochemistry, it can be useful to dilute LowCross-Buffer® in water or physiological buffers. In this case, the user has to determine the ideal mixing ratio by testing.
5. Can buffer components influence the color reaction of enzymatic detection?
No. LowCross-Buffer® does not influence the enzymatic activity of alkaline phosphatase or of peroxidase negatively. Sometimes users detect an increase of enzymatic activity by incubating the labeled detection antibody in LowCross-Buffer®.
6. Can I use LowCross Buffer® in combination with fluorescent dyes, such as in protein arrays?
Yes. For protein arrays there are positive results. Using cyan dyes, such Cy5, Cy3, and Oyster-dyes, may increase fluorescence in some applications.
7. How often can I freeze and thaw LowCross-Buffer®?
LowCross-Buffer® can be frozen and thawed several times, but it is important to mix the buffer thoroughly before use.
8. Can LowCross-Buffer® replace a blocking solution for surfaces?
No. LowCross-Buffer® is not a blocking buffer for surfaces. It can be used for dilution of the specimens or antibodies. For blocking, we recommend The Blocking Solution.