Direct and indirect staining, staining of intracellular antigens, permeabilization and cell preparation protocols
The flow cytometry protocols below provide detailed procedures for the treatment and staining of cells prior to using a flow cytometer.
Protocols are available for:
This is one of the simplest and most common staining methods, where live or fixed cells are incubated with directly labeled antibodies against cell surface antigens
If there is not a directly labeled antibody available, or you wish to amplify your signal you can do indirect staining. This is where you stain a cell with a primary antibody against the antigen of interest and visualize using a labeled secondary antibody which recognizes the primary.
In order to detect antigen not present on the cell surface, cells have to be fixed and permeabilized to disrupt the cell membrane and allow entry of the antibody. Antigens can be then directly or indirectly labeled. Various methods are optimal depending on the antigen and antibody used.
Measuring DNA content for cell cycle analysis requires fixation and permeabilization of the nuclear membrane. We have protocols for staining using DNA binding dyes with and without antibody staining and a protocol for BrdU staining.
Below are protocols for harvesting cells from various sources to obtain healthy cells, essential for optimal staining and analysis.
Specific protocols to use with our antibodies.
General activation protocols using pharmacological reagents and antibodies. Ideal to determine immune competence, marker upregulation, cytokine release and proliferation by flow cytometry.
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