Questions and answers

What is IHC used for?

What is IHC used for?

Immunohistochemistry (IHC) is an important application of monoclonal as well as polyclonal antibodies to determine the tissue distribution of an antigen of interest in health and disease. IHC is widely used for diagnosis of cancers; specific tumor antigens are expressed de novo or up-regulated in certain cancers.

What do you mean by immunofluorescence?

Immunofluorescence (in short, IF) is a method in biology that relies on the use of antibodies chemically labeled with fluorescent dyes to visualize molecules under a light microscope. Schematic of direct (labeled primary antibodies) vs indirect (two antibodies, secondary labeled) immunofluorescence.

What is PBS immunohistochemistry?

Note: PBS buffer is used for routine IHC staining. TBS is used for IHC staining when background is high or when alkaline phosphatase conjugated antibodies are used. PBS is often used for diluting secondary antibodies or streptavidin-HRP conjugate.

How do you Immunostain cells?

Another commonly used immunostaining method is Western blotting where the cells to be analyzed are lysed, and the resulting protein mixture is electrophoresed on a gel to separate the proteins by size. The separated proteins are then transferred to a membrane that can be immunostained.

Why is IHC test done?

IHC, or ImmunoHistoChemistry, is a special staining process performed on fresh or frozen breast cancer tissue removed during biopsy. IHC is used to show whether or not the cancer cells have HER2 receptors and/or hormone receptors on their surface. This information plays a critical role in treatment planning.

How is IHC performed?

Immunohistochemistry (IHC) is a method for detecting antigens or haptens in cells of a tissue section by exploiting the principle of antibodies binding specifically to antigens in biological tissues. The antibody-antigen binding can be visualized in different manners.

What is the principle of immunofluorescence?

An immunofluorescence experiment is based on the following principal steps: Specific antibodies bind to the protein of interest. Fluorescent dyes are coupled to these immune complexes in order to visualize the protein of interest using microscopy.

Why is PBS used in IHC?

This solution helps maintain the morphological characteristics of the antibodies and their respective epitopes, in order to facilitate the specific binding necessary in an IHC reaction.

What is the difference between TBS and PBS?

PBS is Phosphate Buffered Saline which is PH 7.4 and usually used in cell culture, TBS is Tris-Hcl Buffered Saline which is PH7. Also, the phosphate of PBS is more stable in terms of pH vs. temperature than TBS, if you are doing your blocking elsewhere than ambient temperature.

What is immunocytochemistry techniques?

Immunocytochemistry (ICC) is a technique for detection and visualization of proteins, or other antigens, in cells using antibodies specifically recognizing the target of interest. The antibody is directly or indirectly linked to a reporter, such as a fluorophore or enzyme.

What is a drawback of immunocytochemistry?

The disadvantages of IHC are as follows: IHC stains are not standardised worldwide. While the cost of the procedure is relatively inexpensive, the equipment needed to perform IHC is costly. Quantifying results is difficult. IHC is subject to human error.