Scientific Concept of the National Cohort (status ... - Nationale Kohorte

Primary material Volume Processed material
Urine (supernatant) 2.4 ml
Urine (supernatant)
50 ml
Saliva, spitted 2 ml No processing
Stool
Nasal swabs No processing
Small aliquot volumes following
centrifugation
Large aliquot volumes following
centrifugation
(To be determined) minimum
processing, to be elaborared during
feasibility study
119
A.3 Study design
Aliquots and storage
at –180°C
(unless stated otherwise)
12x0.19 ml
5x10 ml aliquots (in
manual –80°C freezers)
2 aliquots (in manual –
80°C freezers)
2 aliquots (in manual –
80°C freezers)
Swab tip (in manual –
80°C freezers)
1 In feasibility studies, both absolute cell recovery and representation of the various mononuclear cell
fractions using Becton Dickinson 'CPT' tubes will be assessed in comparison with conventional Ficoll-
Hypaque gradient centrifugation of acid citrate dextrose (ACD) blood.
All steps of biosample collection, preanalytic handling, storage, and shipping to the central
biobank Table 3.12: will Biomaterials follow specific to be SOPs. collected In addition, in the German the study National personnel Cohort will be trained to ensure
highest and identical quality across all study centers (see also Sect. A.5 on quality assurance
and quality control).
Blood
Blood will be obtained though venipuncture and the procedure carried out by qualified medical
personnel. Even though venipuncture is invasive, it is generally well accepted and any
associated risks can be considered as minimal.
� Overall, 65 ml blood will be obtained from each participant.
� Serum/plasma will be separated from blood cells as quickly and completely as possible
to avoid the occurrence of any artifacts. Serum and plasma will be cooled down
and aliquots prepared that will be stored in the freezer (–80°C) until transport to the
central biobank or storage in the local biobank.
� For hematological analyses, 2 ml EDTA blood will be sent to a certified (clinical) laboratory.
� The buffy coat plus a substantial part of the red blood cell layer (from EDTA tubes)
will be used for isolating peripheral mononuclear cells (PBMC) (after lysis of red
blood cells). PBMC will be resuspended, aliquots prepared, and stored for later DNA
extraction.
� Six aliquots of 0.2 ml packed erythrocytes from EDTA-coated tubes will be stored.
� For each subject, one tube containing RNAse inhibitors will be filled, handled according
to the manufacturer’s instructions, and stored for later RNA extraction. RNA tubes
from PaxGene or Tempus will be used.
� In addition, viable cells will be prepared from a subgroup of participants by means of
Becton Dickinson 'CPT' tubes. For further laboratory work-up, the CPT tubes will be
sent to the laboratory of the central biobank.
A detailed workflow for preparation and storage of locally processed blood and urine samples
has been established. However, some details of this concept are subject to a feasibility
A.3