LowCross-Buffer^® used as an assay buffer helps you to improve signal to noise ratio.
 

Protein array 

ELISA

• Avoid false-positive binding
• Is effective against Human anti-animal antibodies (HAAA), Human anti-mouse antibodies (HAMA), heterophilic antibodies and Rheumatoid Factor
• Decrease CV caused by interferences in human plasma samples
• Eliminate crossreactivities
• Reduce background
• Prevent non-specific binding.
• Eliminate matrix effects

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Frequent asked questions (FAQs):

What is specific to LowCross-Buffer^®?

LowCross-Buffer^® is a newly developed sample buffer for immunoassays (e.g. ELISA, Western blotting, protein arrays and immuno-PCR). It helps to reduce interferences. These are for example, nonspecific binding, cross reactivities and matrix effects. High affinity binding between antibody and antigen is not negatively effected, while unwanted binding of the antibody is inhibited.

Why does LowCross-Buffer^® reduce matrix effects?

Most of the so called matrix effects are based on unwanted low or medium affinity binding of matrix components (e.g. serum proteins from human serum specimen) to analyte or antibody.

The analyte can be masked by proteins or other components of the specimen. Thus the antibody can’t bind the analyte. Also antibodies can be masked by matrix components. LowCross-Buffer^® reduces the masking of antibodies.

How do I have to use LowCross-Buffer^®?

LowCross-Buffer^® is a dilution buffer for specimens (e.g. serum or plasma) and/or for detection antibodies. It depends on the interference. If the nonspecific binding or cross reactivity comes from the detection antibody we recommend diluting the detection antibody in LowCross-Buffer^®.

Do I have to dilute LowCross-Buffer^®?

No, as LowCross-Buffer^® is ready-to-use.

Sample or detection antibodies are diluted directly with the buffer. Only 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 find out the ideal mixing ratio by testing.

Can LowCross-Buffer^® be used with monoclonal and polyclonal antibodies?

Yes, however, if you use polyclonal antibodies they consist of many different antibodies with different affinities. LowCross-Buffer^® only allows high affinity binding whereas low and medium affinity binding will be reduced. If you use LowCross-Buffer^® it can be necessary to increase the concentration of the polyclonal detection antibody moderately. The increase of the antibody concentration also increases the concentration of high affinity antibodies, while LowCross-Buffer^® inhibits binding of low and medium affinity antibodies. Whether you have to increase the concentration of antibodies in the assay or not depends on the quality of the antibodies.

Can buffer components influence the colour reaction of enzymatic detection?

LowCross-Buffer^® doesn’t influence the enzymatic activity of alkaline phosphatase or of peroxidase negatively. Sometimes users detect an increase of enzymatic activity by incubating the labelled detection antibody in LowCross-Buffer^®.

I get a high background in my sandwich ELISA which is detected by a peroxidase labelled secondary antibody. What can be the reason and can LowCross-Buffer^® help?

It’s important to know if the secondary antibody was pre-adsorbed against other antibodies of other mammalian species. Secondary antibodies against rabbit IgG can also crossreact with high affinity with mouse IgG or goat IgG unless purified against antibodies of these species. The reason is the high sequence homology of mammalian antibodies. Secondary antibodies are often not purified (pre-adsorbed) and so crossreact with high affinity. In this case LowCross-Buffer^® will not improve results: replace instead the secondary antibody with a pre-adsorbed antibody which does not crossreact with the capture antibody.

Can I use LowCross Buffer^® in combination with fluorescent dyes e.g. in protein arrays?

Yes, for protein arrays there are positive results. When using cyan dyes (e.g. Cy5, Cy3, Oyster-dyes) an increase of fluorescence in some applications could be observed.

How often can I freeze and thaw LowCross-Buffer^®?

LowCross-Buffer^® tolerates multiple freeze and thaw cycles. Just make sure you the buffer is mixed thoroughly before use.

Can LowCross-Buffer^® replace a blocking solution for surfaces (e.g. Western blotting membranes, ELISA plates)?

No, not at all. LowCross-Buffer^® is not a blocking buffer for surfaces. It is rather an assay buffer to be used for dilution of the specimens or antibodies. For blocking of surfaces such as membranes, we recommend using The Blocking Solution. This casein-based blocker is much more potent than most known blockers commercially available.