Valid

since 28-July-2011 Version 1.0 Page 1 of 3

Working Instruction ABIO-WIS-2011 Blood Pasma and Serum Collection, Processing and Storage
AdeptBio UG (haftungsbeschrnkt) Karl-Wiechert-Allee 76 30625 Hannover, Germany

Table of contents Page 1 Aim of the working instruction ........................................................................................ 2 2 Responsibilities ................................................................................................................ 2 3 Materials and Methods ................................................................................................... 2 4 Disclaimer ........................................................................................................................ 3

Prepared by: Dr. Gabriele Heine Date: 28-July-2011

Approved by: Dr. Michael Jrgens Date: 28-July-2011

Issued by: Dr. Gabriele Heine Date: 28-July-2011

(Signature)

(Signature)

(Signature)


Valid since 28-July-2011 Version 1.0 Page 2 of 3

Working Instruction ABIO-WIS-2011 Blood Pasma and Serum Collection, Processing and Storage
AdeptBio UG (haftungsbeschrnkt) Karl-Wiechert-Allee 76 30625 Hannover, Germany

1 Aim of the working instruction The collection of high quality samples is a prerequisite for research and development in the life sciences industries as it is in academia and should always be carried out under standardized conditions. This instruction describes all steps of collection, processing, storage and transport of blood plasma and serum samples. Blood is collected into serum-gel tubes with coagulation inducer and K2ETDA-containing tubes by venipuncture from a superficial vein of the cubital region to obtain serum as well as plateletpoor plasma. Plasma samples shall be free of haemolysis, coagulation-activation and cellular contamination or platelet activation. Labelling of sample containers has to be performed in a way that allows an unambiguous identification of the sample. 2 Responsibilities AdeptBio provides this working instruction to its partnering collection sites and upon request to all parties who are interested to collect blood serum and plasma samples for research and development under standardized conditions. The collection of blood samples must be performed by authorized medical (laboratory) staff only. 3 Materials and Methods 1. Venipuncture from a cubital vein is performed using a 20 or 21 gauge needle (e.g. 21 g x 0.75 Butterfly Safety-Lok Needle BD # 367281 or similar CE-marked product). If a tourniquet is applied, it should not remain in place for longer than 1 minute (risk of falsifying results due to hemoconcentration). As soon as the blood flows into the container, the tourniquet has to be released at least partially. When more time is required, the tourniquet has to be released so that circulation resumes and normal skin colour returns to the extremity. Aspirate blood into serum containers first (e.g. 9 mL Serum Clot Activator, Gel Separator Vacuette Tube, CE Greiner # 455010 or similar CE-marked product). Depending on size of tube available or volume requested, multiple tubes may be used. Afterwards, blood is drawn into standard K2EDTA containing tubes (e. g. 9.0 ml K2 EDTA Vacuette Tube, CE Greiner # 455036 or similar CE-marked product). Depending on size of tube available or volume requested, multiple tubes may be used. Free flow with mild aspiration has to be assured to avoid haemolysis. Content of tubes is gently mixed by slowly inverting all tubes 10 times. 2. After venipuncture and blood draw: Plasma is obtained from K2EDTA tubes by centrifugation for 10 min at 2,000 x g at room temperature. Preferentially use a centrifuge with swing-out rotor applying no- brake protocols. This centrifugation shall be started immediately after blood


Valid since 28-July-2011 Version 1.0 Page 3 of 3

Working Instruction ABIO-WIS-2011 Blood Pasma and Serum Collection, Processing and Storage
AdeptBio UG (haftungsbeschrnkt) Karl-Wiechert-Allee 76 30625 Hannover, Germany

collection. The resulting plasma sample has now been separated from red and white blood cells in an efficient and gentle way. Nevertheless, a significant number of platelets (~25%) is still present. If platelet-poor plasma is requested an additional step of preparation can be applied as follows. Otherwise proceed and transfer the supernatant in aliquots of 1.0 mL into 2 mL cryovials. Be aware not to touch the buffy coat with the pipet tip during transfer. Optional second centrifugation step for platelet-poor plasma: The plasma sample from the previous centrifugation is transferred into a second vial for another centrifugation for 15 min at 2,500 x g at room temperature. After this centrifugation, the supernatant is transferred in aliquots of 1.0 mL into 2 mL cryovials. Serum is obtained by allowing the Serum tubes to stand upright at room temperature (23-25C) for 30 minutes to allow clotting mechanisms to fully function. Centrifuge at room temperature at 3,000 x g. Preferentially use a centrifuge with swing-out rotor applying no-brake protocols. After centrifugation, the supernatant is transferred in aliquots of 1.0 mL into 2 mL cryovials. 3. Samples are transferred to a 80 C ultrafreezer. Maximum processing time should be under 30 min. for plasma and under 60 min for serum. Storage is at 80C. 4. Transport of samples is performed on dry ice. According to the local situation in hospitals and private practices, adaptations of this working instruction may be necessary. 4 Disclaimer This working instruction is designed to provide optimal blood specimen collection, preparation and storage for general research and development purposes. It was initially developed for analysis of proteins, Peptides and other small molecules present in the fluid. We cannot take responsibility for the usefulness of this working instruction for other purposes. This working instruction is subject to change without prior notice.