BD, 558050, BD Phosflow™ Perm Buffer III

Preparation And Storage: Preparation And Storage Store undiluted at room temperature.
Recommended Assay Procedures: Recommended Assay Procedures Flow cytometry: 1.   Warm BD Phosflow™ Fix Buffer I (Cat. No. 557870) to 37 °C prior to use. 2.   Chill BD Phosflow™ Perm Buffer III to -20 °C prior to use. 3.   Collect PBMCs after Ficoll-Paque separation and treat with any appropriate cell activators as applicable. 4.   At the end of the treatment, immediately mix one volume of the warmed BD Phosflow™ Fix Buffer I with one volume of the PBMC suspension. Mix well and incubate the tubes in a 37 °C water bath for 10 min.  Spin down the cells at 250 x g for 10 min in a table-top centrifuge and then aspirate the supernatant. 5.   Wash the cells once with BD Pharmingen™ Stain Buffer (FBS) (Cat. No. 554656). Alternatively, 1X Washing/Staining Solution (1 x PBS, 1% FCS, and 0.09% sodium azide) may be used. Centrifuge to pellet the cells and remove the supernatant. 6.   Vortex to loosen the cells.  Permeabilize the cells by slowly adding cold BD Phosflow™ Perm Buffer III while vortexing.  Incubate on ice for 30 min and then spin down the cells. 7.   Wash the cells twice with 5-10 mL of BD Pharmingen™ Stain Buffer (FBS). Centrifuge at 250 x g for 10 minutes and remove the supernatant by aspiration. 8.   Resuspend cells in BD Pharmingen™ Stain Buffer (FBS) at 1x 10^7/ml and aliquot 100 µl to each flow tube to continue with antibody staining and flow cytometric analysis. Danger: BD Phosflow™ Perm Buffer III contains 87.68% methanol (w/w). Hazard statements Highly flammable liquid and vapor. Toxic if swallowed, in contact with skin or if inhaled. Causes damage to the central nervous system. Route of exposure: Oral. Precautionary statements Wear protective clothing / eye protection. Wear protective gloves. Take off immediately all contaminated clothing. Wash contaminated clothing before reuse. Wash thoroughly after handling. Use only outdoors or in a well-ventilated area. Keep away from heat/sparks/open flames/hot surfaces. - No smoking. Ground and bond container and receiving equipment.  Use explosion-proof [electrical/ventilating/lighting/] equipment. Use non-sparking tools. Take action to prevent static discharges. Do not breathe mist/vapours/spray. Do not eat, drink or smoke when using this product. IF ON SKIN (or hair): Remove/Take off immediately all contaminated clothing. Rinse skin with water/shower. IF INHALED: Remove victim to fresh air and keep at rest in a position comfortable for breathing. IF SWALLOWED: Immediately call a POISON CENTER/doctor. Rinse mouth. If exposed: Call a poison center/doctor. In case of fire: Use suitable extinguishing media to extinguish. Store in well-ventilated place. Keep cool. Store locked up. Keep container tightly closed.
Product Notices: Product Notices Ficoll-Paque is a trademark of Amersham Biosciences Limited. Alexa Fluor® is a registered trademark of Molecular Probes, Inc., Eugene, OR. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.