Figure 1
UK ME/CFS Biobank procedures from participant recruitment to storage of samples at the UCL/RFH BioBank.
Figure 2
UK ME/CFS Biobank processing procedures conducted by the UCL/RFH BioBank (by tube type with blood derivative breakdowns).
Figure 3
PBMC quality control. PBMC are thawed and stained with either a leukocyte antibody cocktail or a T cell cocktail. To test NK cell function, PBMC are incubated with cytokines at 5 ng/ml for rhIL–12 (Peprotec) and 50 ng/ml for IL–18 (R&D Systems) or with MHC Class I-deficient K562 cells for 18 hours, in the presence of anti-CD107a-FITC (BD Biosciences), then stained with an NK cell function panel. Data are collected using an LSRII flow cytometer using FACS Diva software and analysed using FlowJo software. This figure shows data from samples from 10 participants recruited at different sites throughout 2012. Cell recovery yielded an average of 3.9×10^6 cells/tube. The viability of the recovered cells, assessed by trypan blue exclusion, was excellent in all samples, ranging from 89.2% to 100%. NK cells in the recovered PBMC were responsive in functional NK cell assays, responding to both high-dose cytokines (IL–12 and IL–18) and MHC Class I-deficient (K562) target cells by upregulating expression of the high affinity IL–2 receptor (CD25), increasing production of IFN-γ, and undergoing degranulation, measured by surface expression of CD107a.
Figure 4
PAXgene RNA quality control. RNA is extracted using the PAXgene miRNA isolation kit (Preanalytix). RNA yield is routinely > 3μ g/sample, with optical density 260/280 nm between 1.8 and 2.2 and all Bioanalyser RNA Integrity (RIN) values greater than 7, as expected for intact, non-degraded samples. To confirm samples were appropriate for downstream analysis, quantitative reverse-transcriptase PCR (qRT-PCR) was conducted for five study participants for three housekeeping genes: Human large ribosomal protein P0 (HuP0), cyclophilin A, and hypoxanthine-guanine phospho-ribosyltransferase (HPRT), using published methodology [6].
Table 1
Data available from questionnaires, clinical assessments, and blood tests.
| Questionnaires | Clinical assessments | Blood tests |
|---|---|---|
| • Symptoms experienced | • urinalysis by dipstick (glucose, protein, blood, and specific gravity) | • full blood count |
| • Sociodemographic variables | • pulse oximetry | • blood chemistry and creatinine |
| • Family and individual health histories | • blood pressure (seated and standing) | • liver function |
| • Potential risk factors (exposures) | • standing height | • thyroid function |
| • Medical Outcomes Survey Short Form (SF-36v2) [11] | • weight and bioimpedance | • CRP |
| • General Health Questionnaire (GHQ-28) [12, 13] | • waist circumference | • ESR |
| • Epworth sleepiness score [14, 15] | • hand grip strength test | • rheumatoid factor* |
| • Fatigue scales assessing severity [16, 17, 18] and disability [17, 19] | • spirometry | • tissue transglutaminase antibodies* |
| • Pain and fatigue analogue scale [20, 21] | • serum vitamin B12* | |
| • folate* |
[i] * Available for baseline only.
Table 2
UK ME/CFS Biobank sample holdings at the UCL/RFH BioBank (through October 2016).
| Sample type | Stored aliquot/tube volume | Collection tube (BD) | Number of aliquots |
|---|---|---|---|
| Whole blood | 0.5 ml | EDTA | 3,021 |
| Serum | 200 μl | Red serum | 6,770 |
| Plasma | 1 ml | EDTA | 2,029 |
| Plasma | 250 μl | Na Hep | 5,382 |
| Red Blood Cells (RBCs) | 2 ml | Na Hep | 773 |
| Peripheral Blood Mononuclear Cells (PBMCs) | 5×106 cells in 1 ml | Na Hep | 9,106 |
| Peripheral Blood Mononuclear Cells (PBMCs) | 5×106 cells in 1 ml | EDTA | 2,198 |
| Blood for RNA | 2.5 ml | PAXgene | 584 |
| Total | 29,863 |
Figure 5
Participant invitation and enrolment figures (through October 2016).
Figure 6
Workflow for accessing samples and/or data.