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This ready to use Negative Control reagent contains purified, fluorescein or phycoerythrin conjugated mouse immunoglobulin molecules of IgG1 isotype, which have been selected on the basis of their binding characteristics: no specific binding to human cell surface or intracellular antigens, same low range of nonspecific binding to human leukocytes as other Reagents. This isotype control IgG1 is suitable as a Negative Control to be used in combination with reagents for the: - Enumeration of Myeloid Cells - Analysis of Myeloid Differentiation Stage - Enumeration of B-cells and Precursors - Enumeration of T-cells and Precursors - Analysis of Leukemia Cells - Analysis of Immunodeficiency StatesThe negative control reagent permits to estimate the degree of non-specific binding of isotype matched immunologbulins to leukocytes via e.g. Fc-receptors. It enables the expert to set flow cytometric parameters accordingly.Results must be put within the context of other diagnostic tests as well as the clinical history of the patient by a certified professional before final interpretation.
Procedure: For each sample to be analysed use 50 µL of whole blood, bone marrow or mononuclear cell suspension in a 5 mL tube Add 100 µL of Reagent A (Fixation Medium, stored and used at room temperature) Incubate for 15 minutes at room temperature Add 5 mL phosphate buffered saline and centrifuge cells for 5 minutes at 300 g Remove supernatant and add to cell pellet 100 µL Reagent B (Permeabilization Medium) and 20 µL of the appropriate monoclonal antibody conjugate Vortex at low speed for 1-2 seconds Incubate for 15 minutes at room temperature Wash cells with phosphate buffered saline as described above Remove supernatant and re-suspend cells in sheath fluid for immediate analysis or re-suspend cells in 0.5 mL 1.0 % formaldehyde and store at 2-8 °C in the dark Analyse fixed cells within 24 hours