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UNIVERSITY OF LEICESTER CANCER STUDIES & MOLECULAR MEDICINE STANDARD OPERATING PROCEDURE SOP - 635
VERSION - 01
TITLE: Receipt of whole blood and isolation of plasma and buffy coat for melanoma research projects (with approved ethics and consent) Written by: Linda Potter Reviewed by: Angie Gillies Changes introduced:
Implementation date: 12.4.12 Review date: 12.4.14
1. PURPOSE To receive whole blood (with consent) and isolate plasma and buffy coat.
2. SPECIAL NOTES Full details of the use of this procedure must be recorded in the appropriate laboratory notebook, in accordance with SOP 105.
3. CROSS REFERENCES 3.1 3.2 3.3
Maintenance of laboratory note book RNA/DNA extraction from whole blood Melanoma blood sample collection book
Jouan centrifuge Class 1 cabinet room 342B 15ml falcon tubes 1.5ml eppendorf tubes P1000 gilson and 1ml tips Tri-gene SLS 703725-1 Plastic box with lid for disposal of contaminated tubes, tips etc.
500 series\sop635v01
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5. PROTOCOL Blood samples are to be collected from Clinic 3 in the Balmoral building every Monday and Friday between 8.30am and 1pm. At 8am take a lidded polystyrene box containing ice to the clinic. Any blood samples taken will be kept on ice until collected. Check at 11am and 1pm and bring blood samples to the Lab. Check the details and the consent form. Book in using the H number system: In the H book write the unit number and the patient’s surname in pencil. Write the H number and the sample type (pre-op or post-op) on the consent form. Record the details in the melanoma blood sample collection book. Write the H number on the blood sample. Aliquots of the isolated plasma will be given the H number and the buffy coat will be given the H number followed by BC.
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Transfer fresh blood to a 15ml falcon tube labelled with the H number. Spin fresh blood in EDTA tubes, in Jouan centrifuge at 2000 rpm/850 g for 10 minutes at 4°C. In the Class 1 cabinet, remove plasma and decant into 15ml falcon tube. Remove buffy coat and decant into an eppendorf tube. Spin plasma in Jouan centrifuge at 2000 rpm/850 g for 10 minutes at 4°C. Remove plasma (leaving cell debris in tube for disposal) and decant into 1ml aliquots in eppendorf tubes. Re-spin plasma in microfuge at 13500 rpm/20 000g for 5 minutes at room temp. Decant into 1ml aliquots in eppendorf tubes. Freeze all plasma and buffy coat aliquots at -80C and store in room 302. Keep a record of the number of aliquots of plasma and buffy coat, which box they are stored in and the freezer location. Write this information on the consent form under the H number. Place all contaminated tubes, tips etc. in diluted Tri-gene (1:100). Leave overnight and dispose of in the clinical waste in room 320. Update the blood samples spreadsheet and file the consent forms.