A Brief at the Variant Flow Cytometry Antibody Staining Procedure

Posted by Facs Analysis on September 6th, 2019

Flow cytometry is the latest laser-based technology to evaluate the expression of cell surface and intracellular molecules and determining the varying cell types in a heterogeneous cell population. It also assists in accessing the purity of isolated subpopulations and specifying the cell size and volume. Besides, FC is used to analyze the fluorescence intensity given by fluorescent-labelled antibodies.

The General Experimental Procedure of staining comprises making a single-cell stay from cell culture or tissue samples. Afterwards, the cells then incubated in tubes with unlabeled or fluorochrome-labelled antibodies and evaluated on the flow cytometer.

Direct staining:

During the straightforward Flow Cytometry Antibody Staining Procedure, cells are incubated with an antibody straightly conjugated to a fluorochrome. It needs to require only one antibody incubation step and deducts the probability of non-specific binding from a lower-level antibody.

This step is required for intracellular staining, in which large antibody-fluorochrome complexes comprise the low-level antibodies can become confined, resulting in interrupting to enter the cell halting the primary antibody detection.

Intracellular staining:

Flow Cytometry Antibody Staining Procedure of intracellular antigens relies on variant fixation and permeabilization strategies to permit access of antibodies to internal cellular proteins. An assured staining process in all cases is dependent on the optimization of experimental situations with the help of titering of antibodies, usage of suitable controls to installation the FC efficaciously and optimized fixation and permeabilization process.

Indirect staining:

In indirect staining, the primary antibody is not fluorochrome-labelled; however, it is detected by a fluorochrome-labelled low-level antibody. This second reagent can be an antibody with specificity for the primary antibody. Also, the avidin-biotin system is taken into consideration, whereby an antibody is conjugated to biotin and detected with fluorochrome-labelledavidin.

Detection of secreted proteins:

To deduct the secret protein, it is a daunting process as the protein will be coming from the before detection. Moreover, a Brefeldin A may be used to inhibit secretion of expressed proteins. Afterwards, the intracellular staining method will be used to deduct the target protein.

If you need to take information about the FACS Antibody, then you need to log in Booster Antibody and Alisa Experts atfacs-analysis.com. You will able to find out the experts over there, which will assist you to give the information required for the general experimental procedure. You can contact this destination through email or over the phone.