Phospho Antibodies

What Are Phospho Antibodies?

Protein phosphorylation is a process where a phosphate group is added to certain amino acid residues of a protein – usually serine (S), threonine (T), or tyrosine (Y) - by enzymes called kinases. This process is integral in controlling cellular signaling, cellular growth, and other biological functions.

Our catalog includes a wide range of phospho-specific antibodies that can accurately detect this important marker. They perform strongly in widely-used laboratory applications such as Western blot, flow cytometry, immunohistochemistry, and immunofluorescence microscopy. We value your trust in us and are committed to providing top-quality products and services. All of our antibodies are guaranteed to work for the applications and species indicated on our website & associated product pages.

What Are The Key Applications of Phospho Antibodies?

1. Western Blotting

One of the first steps a researcher can take in utilizing these phospho-specific antibodies, is to check if the antibody works using a technique referred to as “Western blot”. For those unfamiliar, Western Blot aids in showing whether the protein that the antibody recognizes is appearing at the correct/expected size. These phospho-specific antibodies should also be able to detect changes in the target protein’s phosphorylation (on/off state) when cells are stimulated in certain ways.

2. Staining of Fixed Cells (Immunocytochemistry)

Another routine use of these phospho-specific antibodies, is to test if the antibody is able to demonstrate similar performance when used on fixed cells (intact cells that have been preserved) as it did in the Western blot tests. It is an important aspect in many cases to confirm that the antibody works in actual intact cell samples. Ideally, the method used for cellular fixation should be the same as what is used in pathology labs (like using 10% formalin). To check if the antibody works well in tissue sections (FFPE), researchers will often test it on fixed cells that are processed similar to tissue samples.

3. Specificity Tests Using Peptides

In order to make sure that the antibody is only binding to the right target:

• Laboratory technicians will mix the antibody with phospho-peptides (short segments of the protein containing the phosphate group modification).


• If the antibody signal disappears, it is confirmation that it is binding to the correct phosphorylated location.


• A more robust test is to use both the phosphorylated and non-phosphorylated (dephosphorylated) versions of the protein. The antibody should react only with the phosphorylated one.


• Another method sometimes utilized is to treat the sample with an enzyme, such as alkaline phosphatase, that specifically removes phosphate groups. If the antibody signal disappears after this, it also confirms specificity.

4. Genetic Confirmation

As a final step, scientists can genetically manipulate the nucleotide sequence and alter the target protein by removing the exact site where phosphorylation happens. If the antibody no longer appears to detect the modified protein, it is strong evidence supporting the antibody being specific for that phosphorylated site.

Why Buy Phospho Antibodies Through Us?

• The production laboratory adheres to strict and consistent protocols prior to releasing any of these phospho-specific antibodies:


• Standard methods and proper controls in all tests to ensure high quality.


• These antibodies are tested and validated in different cell types and species.


• High quality control criterion to ensure each batch is consistent, so you will obtain reliable results every time.

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