TransFix Cellular Antigen Stabilising Reagent C€ ivd Safety Data Sheet can be obtained at www.caltag.co.uk Intended Purpose TransFix® and TransFix Sample Storage Tubes are intended for stabilisation and storage (in the case of Sample Storage Tubes) of whole blood specimens for immunophenotyping of white blood cells by flow cytometry. Recovery of lymphocyte subset markers can be accomplished over a 14-day period following collection. TransFix is for In-Vitro Diagnostic Use. Immunophenotyping by flow cytometry provides a rapid and accurate assessment of the frequency and type of leucocytes in a blood sample. Any delay in testing the sample, such as transport from the collection site to analysis location, can have a negative impact on results. Sample preservation provides a solution that addresses delays between sample collection and testing. Subsets of leucocytes can be distinguished by cell surface antigens using fluorescent antibodies and flow cytometry. Qualitative and quantitative changes in leucocyte subsets are used to identify and monitor immunodeficiency and haematological diseases [1]. The TransFix stabiliser acts by preserving the cell surface antigens of lymphocyte subsets until processing and analysis can be performed. TransFix is available in 1 mL and 20 mL aliquots in polypropylene vials, or as a 'sample storage tube': consisting of a 2 mL polypropylene vial containing 0.2 mL TransFix. 1. TransFix is intended for use as specified in this document. It is an in-vitro diagnostic product for professional use only. 2. TransFix and TransFix Sample Storage Tubes are only for use with flow cytometry analysis. 3. Anti-coagulated blood must be kept at room temperature prior to treatment with TransFix. 4. Incubation times or temperatures other than those specified will lead to erroneous results. 5. Do not use TransFix after the expiration date on the tubes and packaging. 6. Do not dilute or add other components to TransFix. 7. Do not use cell viability stains on samples treated with TransFix as they are fixed instantaneously. 8. TransFix Sample Storage Tubes are single-use only. Do not re-use. 9. Ensure the correct sample to TransFix ratio is used as per instructions; an incorrect blood-to-additive ratio and may lead to poor product performance and incorrect analytic results. 10. TransFix treated samples and all materials coming into contact with it should be handled as if capable of transmitting infection. TransFix is a clear green liquid containing formaldehyde and other chemicals. H317 - May cause an allergic skin reaction. P261 - Avoid breathing fume, mist, spray, vapours. P280 - Wear protective gloves, protective clothing, eye protection. P302+P352 - IF ON SKIN: Wash with plenty of soap and water. P308+P313 - IF exposed or concerned: Get medical advice/attention. P362+P364 - Take off contaminated clothing and wash it before reuse. Indications of Product Deterioration 1. Cloudiness or precipitate visible in the TransFix. 2. Colour change of TransFix from a clear green liquid. 3. TransFix changes from liquid to solid. If indications of product deterioration occur, do not use and contact Caltag on: +44(0)1280 827460 or [email protected]. TransFix and TransFix Sample Storage Tubes should be disposed of in accordance with local regulations. Avoid disposing into drainage systems and the environment. All TransFix products are shipped at ambient temperature (2 - 25°C). Additional insulation may be required for shipment during extreme temperature conditions. Store TransFix and TransFix Sample Storage Tubes at 2 - 25 °C before use. Materials needed but not supplied • Blood collection tubes and materials for sampling. • Automatic pipettes with disposable tips for 10, 20, 100 and 1,000 pL. • Flow cytometry tubes • Flow cytometry calibration and control reagents • Specific fluorescent antibodies and isotype controls • Red blood cell lysing solution • Automatic agitator/vortex machine • Flow cytometer 1. Collect blood by venepuncture into an EDTA vacuum tube according to normal phlebotomy practise. Treat sample with TransFix within 6 hours of venepuncture: 2. For TransFix Sample Storage Tubes, mix the blood sample, remove 1 mL and add to the tube. 3. For TransFix reagent, carefully remove the blood collection tube cap and measure the volume of anti-coagulated whole blood within the vacuum tube. Pipette into the blood collection tube the appropriate volume of TransFix at the ratio of 0.2 mL TransFix per 1 mL of blood. 4. Replace the cap on the sample tube, ensuring that there is no leakage. 5. Mix the treated sample carefully. Do not vortex. 6. Store / transport the TransFix treated blood for up to 14 days at 2 - 8 °C, Store / transport the TransFix treated blood at 18-25 °C for up to 3 days, and at 2-8°C for a up to a further 4 days. Incubate the treated sample at room temperature (18-25 °C) for 15 minutes before use. 7. Re-suspend the cells thoroughly by gently inverting the tube at least 10 times. Heavier cells and blood components will sediment over time, forming two distinct layers; this is normal. 8. Take care when opening TransFix Sample Storage Tubes as there may be a build-up of blood in the caps. 9. Stain and analyse your stabilised blood sample as per your usual flow cytometry protocol (N.B. see Notes). 1. It is recommended that all antibody conjugates are validated in association with TransFix prior to use. 2. Use caution when implementing automatic gating strategies as light scatter positions of cells stabilised by TransFix may differ from those of untreated cells. 3. Gating for lymphocytes should be performed on CD45+/SSc. 4. If calculating absolute cell counts, the dilution with TransFix must be accounted for. I.e., adjust the absolute cell count by multiplying the output by 1.2. Expected Results Immunophenotyping of blood samples stabilised with TransFix for 14 days are comparable to fresh samples. See Table 1 for example data.