Dear Suzanne,

In general, buffers provide a stable condition regarding the pH during a reaction. Every protein and enzymes require specific pH and buffer condition for their normal structure and optimal function. Without a stable bufferic environment, fluctuations in pH can disturb the normal function of different elements in a biological procedure including biochemical interactions. This stable condition is crucial for ELISA, since the binding of antibody to target protein is highly sensitive to pH, even binding of blocking reagent and secondary antibody, also the function of peroxidase enzyme. Similarly, in DNA extraction via column, the binding of DNA to column is also based on ionic interactions, which are sensitive to pH too.

Hi Suzanne, physiological buffer solutions are vital for most protein structure and function.

Dear Suzanne

ELISA protein extraction buffer contains a mild detergent enabling desolution of cell membranes, solubilization of most membrane proteins and suspension of cytosolic proteins into the buffer.

Dear Seied, thank you for your informative answer.

Hi Steingrimur, what do you mean of vital for protein structure? do you mean to maintain protein structure? do you mean changing in PH may disrupt the protein structure thus the protein function?

Dear Seied , in ELISA technique we deal with serum in most cases and as you know there are no cells in serum to destroy their membrane, and what is the benefit of the cytosolic protein being suspended? can you give me some ideas, please.

Dear Suzanne

Several different buffers are used during an ELISA: one for coating, another for blocking, another for washing, and perhaps another for sample and antibody dilution. Some specialist buffers are multifunctional, allowing for simultaneous coating, stabilization, and blocking. One such example is General Assay Diluent (BUF037), which is suitable for most sandwich ELISAs and formulated for use with serum and plasma samples. It contains goat sera to reduce non-specific interactions and a proprietary calcium chelating agent to inhibit interference from thrombin and complement.

https://www.bio-rad-antibodies.com/elisas-buffers-substrates.html

https://www.bio-rad-antibodies.com/helpful-elisa-hints.html

Dear Suzanne,

In general, buffers provide a stable condition regarding the pH during a reaction. Every protein and enzymes require specific pH and buffer condition for their normal structure and optimal function. Without a stable bufferic environment, fluctuations in pH can disturb the normal function of different elements in a biological procedure including biochemical interactions. This stable condition is crucial for ELISA, since the binding of antibody to target protein is highly sensitive to pH, even binding of blocking reagent and secondary antibody, also the function of peroxidase enzyme. Similarly, in DNA extraction via column, the binding of DNA to column is also based on ionic interactions, which are sensitive to pH too.

Dear Amir, thanks a lot