BD Pharmingen™ PE Rat Anti-Human IL-6

Immunofluorescent Staining and Flow Cytometric Analysis: The MQ2-6A3 antibody is useful for immunoflourescent staining and flow cytometric analysis to identify and enumerate human IL-6 producing cells within mixed cell populations. This 100 Test Size formulation of the PE-conjugated MQ2-6A3 antibody has been pre-titrated to assure effective intracellular detection of human IL-6 using 20 µl/1 x 10e6 cells.

A useful control for demonstrating specificity of staining is either of the following: 1) pre-block the conjugated MQ2-6A3 antibody with a ligand (e.g., recombinant human IL-6; Cat. No. 550071) prior to staining, or 2) pre-block the fixed/permeabilized cells with unlabelled MQ2-6A3 antibody (Cat. No. 559068) prior to staining. The staining technique and blocking controls are described in detail by C. Prussin and D. Metcalfe. A suitable rat IgG2a isotype control for assessing the level of background staining on paraformaldehyde-fixed/saponin-permeabilized human cells is also available in a 100 Test Size formulation PE-R35-95 (Cat. No. 559317).

Important Note: This pre-titered antibody solution does not contain a cell permeabilization agent. It is necessary to include a cell permeabilization agent when using the pre-titered antibody solution to stain fixed and permeabilized cells. Perm/Wash™ Buffer (Cat. No. 554723) contains the permeabilization agent saponin and is useful for this purpose as described in the USAGE section below.

USAGE

1. Resuspend 1 x 10e6 fixed and permeabilized cells in 20 µl of the pre-titered antibody solution and 30 µl of 1X Perm/Wash™ Buffer (Cat. No. 554723).

2. Incubate the cell suspension for 15 minutes (at RT or 4°C).

3. Wash twice in 100 µl of 1X Perm/Wash™ Buffer (Cat. No. 554723).

1. This reagent has been pre-diluted for use at the recommended Volume per Test. We typically use 1 × 10^6 cells in a 100-µl experimental sample (a test).
2. Since applications vary, each investigator should titrate the reagent to obtain optimal results.
3. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
4. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
5. Caution: Sodium azide yields highly toxic hydrazoic acid under acidic conditions. Dilute azide compounds in running water before discarding to avoid accumulation of potentially explosive deposits in plumbing.
6. Source of all serum proteins is from USDA inspected abattoirs located in the United States.