FIX&PERM® Cell Fixation and Permeabilization Kit (CE)

Nordic Mubio
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Product Name FIX&PERM® Cell Fixation and Permeabilization Kit (CE)
Description This FIX&PERM® Cell Fixation and Permeabilization Kit contains 2 reagents: Fixation Medium (Reagent A) and Permeabilization Medium (Reagent B). It is intended for first fixing cells in suspension with Reagent A and then permeabilizing the cell membranes with Reagent B. This procedure gives antibodies access to intracellular structures and leaves the morphological scatter characteristics of cells intact. Specific formulations reduce background staining and allow simultaneous addition of permeabilization medium and fluorochrome labeled antibodies. FIX&PERM® is suitable for the analysis of normal and malignant leukocyte populations derived from various human biological samples (blood, bone marrow and others) using flow cytometry. 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. Format: 4 x 5ml vial of Fixation Medium (Reagent A) and 4 x 5ml vial Permeabilization Medium (Reagent B). FIX&PERM® Reagents are designed for use with all commercially available flow cytometers. Alignment and compensation should be performed according to manufacturer´s instructions. The quality of each FIX&PERM® Lot is determined by fixation and permeabilization of well defined blood samples from representative donors and subsequent comparison of forward and side scatter characteristics of obtained leukocytes, as well as immunostaining efficiency for several membranous and cytoplasmic antigens
Applications -, And others) must be collected under sterile conditions. Anticoagulation with EDTA or heparin is recommended. The samples should be stored at room temperature until used. For optimal results, Biological fluids (blood, Bone marrow, Bone marrow or mononuclear cell suspension in a 5ml tube €¢ Add 100 µl of Reagent A (Fixation Medium, Etc., Fixation, Lab-coat, Permeabilization and staining procedure:, Samples should be processed and analyzed within 24 hours. Samples with high numbers of non-viable cells might cause false results, Stored and used at room temperature), Such cases require determination of cell viability with e.g. propidium iodide. All biological samples have to be handled with caution. Always consider them as potentially infective. Use appropriate precautions such as gloves, €¢ Add 5ml phosphate buffered saline and centrifuge cells for 5 minutes at 300 g, €¢ For each sample to be analyzed add 50 µl of whole blood, €¢ Incubate for 15 minutes at room temperature, €¢ Remove supernatant and add to cell pellet 100 µl Reagent B (Permeabilization Medium) and 20 µl of the appropriate Nordic-MUbio monoclonal antibody conjugate, €¢ Remove supernatant and resuspend cells in sheath fluid for immediate analysis or resuspend cells in 0.5 ml 1.0 % formaldehyde and store them at 2-8C in the dark. Analyze fixed cells within 24 hours., €¢ Vortex at low speed for 1-2 seconds, €¢ Wash cells with phosphate buffered saline as described above
Storage FIX&PERM® Cell Fixation and Permeabilization Kit reagents should be stored and used at room temperature. Do not freeze. Stability of the reagent: Please refer to the expiry date printed onto the vial. The use of the reagent after the expiration date is not recommended. If reagents are stored under any conditions other than those specified, the conditions must be verified by the user. Do not use reagents if a precipitate should form or discoloration occurs. If unexpected results are obtained which cannot be attributed to differences in laboratory procedures, please contact us.
References - Gerna, G., Percivalle, E., Lilleri, D., Lozza, L., Fornara, C., Hahn, G., Baldanti, F. & Revello, M. G. (2005) J Gen Virol 86, 275-84.
- Groeneveld, K., te Marvelde, J. G., van den Beemd, M. W., Hooijkaas, H. & van Dongen, J. J. (1996) Leukemia 10, 13
Limitations Flow cytometry should be performed by professional users only. Improper alignment of the flow cytometer, inaccurate compensation of fluorescence leaking into other channels as well as incorrect positioning of regions may lead to false results. Lysis of red cells might be impossible for various reasons. In such instances it is recommended to isolate mononuclear cells (MNC) via density gradient centrifugation prior to staining. Results will be correct and reproducible as long as the procedures used respect the technical recommendations and obey good laboratory practice. The FIX&PERM® solutions are provided in a concentration that will allow to fix and permeabilize human hematopoietic cells. It is therefore strongly recommended to stick to the working protocol in terms of concentration and volume regarding cells and antibody. The properties of FIX&PERM® have been determined using EDTA anti-coagulated peripheral blood.

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