Biosample quality for OMICs downstreamanalysis

Gabriele AntonMilan, 2015

Standardization of Sample Handling

Literature review

Best practices of

participating cohorts

and experts

within the consortium

Scientific research

Life cycle of a biospecimen

Végvári A, Welinder C, Lindberg H, Fehniger TE, Marko-Varga G - (2011)

The patient

Fasting vs. non-fasting

The patient – fasting status

Variation in metabolites

during fasting and subsequent

SLD (standard liquid diet)

+ interindividual variation

Krug et al. 2012

The patient

Fasting vs. non-fasting

Sex, age, BMI

Lifestyle (long-term, short-term prior to sampling,

e.g heavy physical activity, alcohol consumption)

The patient – alcohol consumption

Same metabolites affected

in atherosclerosis,

inflammatory diseases

or ischaemia → correction

necessary

Jaremek et al. 2013

The patient

Fasting vs. non-fasting

Sex, age, BMI

Lifestyle (long-term, short-term prior to sampling,

e.g heavy physical activity, alcohol consumption)

Pregnancy

Menstrual cycle

Medication (e.g. surgery patients)

Documentation!!!

Sampling

Decision on matrix (e.g. plasma vs. serum)

Sampling

Yu et al, 2011

Concentration differences

in metabolites

between plasma and serum

samples of the same probands

Sampling

Decision on matrix (e.g. plasma vs. serum)

Position of the proband, resting time before sampling,

Clamping, manipulations (warmth, pumping) (blood)

Location for sampling (tissue specificity of e.g. epigenetic marks)

Disinfection of sampling site

Sampling - disinfection

Van der Sar et al, 2015

Variable

Ethanol signal

in CSF samples

measured by

H-NMR

metabolomics

Sampling

Position of the proband, resting time before sampling,

Clamping, manipulations (warmth, pumping) (blood)

Location for sampling (tissue specificity of e.g. epigenetic marks)

Disinfection of sampling site

Collection devices, additives, order of tubes (tubes with additives last)

Sampling – device

Breier et al, 2014

Difference in Methionine

Sulfoxide between serum

tubes with clotting

activator and gel-barrier tubes

Sampling

Position of the proband, resting time before sampling,

Clamping, manipulations (warmth, pumping) (blood)

Location for sampling (tissue specificity of e.g. epigenetic marks)

Disinfection of sampling site

Collection devices, additives, order of tubes (tubes with additives last)

Standardization and Documentation!!!

Sample processing

Time:

Blood, urine: until centrifugation cells still in contact with liquid, artefacts

by cell rupture; avoid harsh treatment like shaking, low temperatures)

Processing after centrifugation: cooling, fast sample handling

Sample processing – Time/Temperature

Breier et al, 2014

Changes in plasma

metabolites due to

handling time

and temperature

Sample processing – Time/Temperature

Ratio of total

lysophosphatidylcholines to

total phosphatidylcholines

in serum samples stored at

different temperatures

Anton et al, 2015

Sample processing – Time/Temperature

Anton et al, 2015

Marker for prolonged

room temperature

exposure

Sample processing

Time:

Blood, urine: until centrifugation cells still in contact with liquid, artefacts

by cell rupture; avoid harsh treatment like shaking, low temperatures)

Processing after centrifugation: cooling, fast sample handling

Tissue: warm and cold ischemia time, type of conservation

Standardization and Documentation!!!

Sample processing

Processing requirements depend on desired downstream analysis:

DNA quite stable (but: epigenetics?),

proteins and metabolites very variable stability

RNA rather instable

For biobanked samples markers can help to take decisions on

possible downstream applications

Sample Transport

TimeTemperatureUse loggers to document conditions

Documentation!!!

Sample Storage

Liquid samples: small aliquots to avoid freeze-thaw

Temperature: liquid nitrogen vs. -80°C ?

New solutions for room temperature storage of e.g DNA

Summary

Use standardization as far as possible, especially

for multi-center studies

Good documentation of all steps of biosample generation

very important (SPREC)

For biobanked samples, markers for desired downstream

analysis necessary (quality control)

Acknowledgements

The research leading to these results has received funding from the European Union Seventh Framework Programme (FP7/2007-2013) under grant agreement n° 261433 (Biobank Standardisation and Harmonisation for Research Excellence in the European Union - BioSHaRE-EU)

Participating biobanks, especially: Life Lines - UMGC, HUNT, KORA, German National Cohort and their staff

BioSHaRE collaborators

Questions?

Gabriele Anton

Research Unit of Molecular Epidemiology (AME)Institute of Epidemiology IIGerman Research Center for Environmental Health

gabriele.anton@helmholtz-muenchen.dehttp://www.helmholtz-muenchen.de/ame