Biological samples to be collected

There was extensive consultation and discussion on which biological samples to collect at the assessment centre visit. The inclusion criteria were based on the likely value of the additional information that would be made available by collecting some particular sample type (i.e. the range of assays that could be made and the physiological coverage of the material), and the feasibility and cost of collecting and processing such samples from the 500,000 participants.

On this basis, it was decided to collect 40-50 ml of blood and a random urine sample during the baseline assessment visit (see Box 1.5.1).

Sample type Selection criteria

Blood

• Variety of fractions: plasma, serum, white cells, red cells, peripheral blood lymphocytes

• Wide range of biomolecules: DNA, RNA [5’ ends], proteins, analytes

• Wide physiological coverage: genome, proteome and metabolome, haematological parameters

• Suitable for a very wide range of assay technology

• Ease and low cost of collection

Urine

• Wide range of biomolecules: proteins, analytes (including pharmaceuticals)

• Wide physiological coverage: proteome and metabolome (including gut microbiome)

• Suitable for many assay/technology types

• Low cost of collection

Box 1.5.1: Included biological samples and rationale

Having decided on blood and urine collection, consideration was given to additional types of sample that might allow measurements of factors not covered by blood or urine (see Box 1.5.2). On this basis, it was decided to exclude all other sample types because they were not considered likely to provide sufficient additional information to characterise participants in ways that would be importantly predictive of subsequent health outcomes. For example, bacterial gut fermentation by-products in faeces are biomarkers of a number of diseases of the gut (such as irritable bowel syndrome and, possibly, Crohn’s disease). These markers include hydrogen, methane, alkanes, methyl alkanes, phenols and organic acids, which can also be measured accurately in urine [106]. Furthermore, the gut microbiome can be profiled in urine using NMR approaches. Hair and nails may be used to assess medium-term exposure to heavy metals. But, a study of the toxicokinetics of methylmercury exposure concluded that hair and blood levels are of questionable value as indicators of both body and target organ concentrations of mercury [107]. Moreover, some forms of arsenic (such as arsenobetaine, the major organic arsenic compound in seafood) do not accumulate in hair [108]. In addition, measures of environmental arsenic in hair and nails are influenced by external contaminants (such as air, water soaps and shampoos), and such exposure is better measured in urine [109].

Sample type Exclusion criteria Faeces

• Limited additional information (e.g. gut microbiome )

• Difficulty in collecting/processing

• Potential impact on recruitment

• Complexity and cost of storage Hair

• Limited additional information (e.g. exposure to environmental heavy metals )

• Complicating effects of cosmetics and toiletries

Nails

• Limited additional information (e.g. exposure to environmental heavy metals )

• Complicating effects of cosmetic products

• Inconsistency of sample collection

• Possible impact on recruitment of clipping nails Saliva

• Limited additional information (e.g. indicators of periodontal disease and oral cancer)

• Extra cost of storage

Box 1.5.2: Excluded biological samples and rationale 1.5.3 Types of sample collection tubes

There is a very wide variety of preservatives and additives available for the collection of blood and urine. In a review of factors that affect the quality of biomarker assays, the importance of careful selection of anticoagulants and preservatives in the collection tubes was stressed [110]. Certain anticoagulants are recommended for some analyses whilst others are contraindicated. For example, blood collected into EDTA-containing tubes is good for DNA-based assays, but may be unsuitable for others because it

chelates magnesium ions; heparin-stabilized blood affects T-cell proliferation assays and heparin binds to many proteins. EDTA plasma and serum give assay-dependent variation in measures of growth hormone, thyroid stimulating hormone, insulin, C-peptide, total estradiol, testosterone, cortisol and progesterone in fluorometric and immunofluorometric assays [111]. Any anticoagulant may cause in vitro induction of cytokines and artefactually elevated concentrations [112]; and addition of borate stabilises urine samples but interferes with some metabonomic assays (Jeremy Nicholson: personal communication). Inevitably, the selection of additives is a compromise, and the choice made for UK Biobank has been made to cover as wide a range of potential future uses as is feasible.

UK Biobank’s sample handling pilot studies have demonstrated that maintaining whole blood and urine samples at 4^oC for at least 36 hours prior to processing and cryopreservation allows a very wide range of assays to be performed [105]. An additional acid citrate dextrose (ACD) tube of whole blood maintained at 18^oC also allows subsequent immortalisation of lymphocytes. Consequently, the processing of collection tubes at the assessment centre can be minimised, and most of the processing conducted at the central laboratory using efficient automated systems. These processing platforms isolate and aliquot multiple fractions from the EDTA tubes to produce fractions suitable for DNA extraction and a wide range of assays using the red cells and plasma. In addition, one gel plasma separation tube (PST) and one gel serum separation tube (SST) will be collected for each participant to protect the plasma/serum from any changes prior to delayed separation that might affect certain assays (e.g. elevation in the levels of potassium and homocysteine).

Type of sample Collection priority

Table 1.5.1: Sample collection priority, volume and transfer temperature The “vacutainer” system will be used to collect these blood and urine samples (see Table 1.5.1). During venepuncture, the hypodermic needle is connected to these vacutainer tubes, which are held under a slight vacuum and contain the required additives, and the vacuum draws sufficient blood to fill them. As a set of the required tubes is collected, unique bar-codes for each tube are scanned into the assessment centre IT system to link each tube with the participant’s identifier number. A collection priority is specified in the event that assessment centre staff cannot extract sufficient blood for the full set of tubes in order to provide the widest possible range of different fractions and

the participant’s urine into a vacutainer from the urine collection vessel. All tubes are maintained at 4^oC (with the exception of the ACD tube which is maintained at 18^oC) until they are ready for dispatch to the central processing laboratories in temperature-controlled shipping boxes.