General DBS procedure

Review

Sylvain Lehmann*, Constance Delaby, Jérôme Vialaret, Jacques Ducos and Christophe Hirtz

Current and future use of “dried blood spot”

analyses in clinical chemistry

Abstract: The analysis of blood spotted and dried on a matrix (i.e., “dried blood spot” or DBS) has been used since the 1960s in clinical chemistry; mostly for neona- tal screening. Since then, many clinical analytes, includ- ing nucleic acids, small molecules and lipids, have been successfully measured using DBS. Although this pre- analytical approach represents an interesting alternative to classical venous blood sampling, its routine use is lim- ited. Here, we review the application of DBS technology in clinical chemistry, and evaluate its future role supported by new analytical methods such as mass spectrometry.

Keywords: dry blood spot; enzyme-linked immunosorb- ent assay (ELISA); mass spectrometry; polymerase chain reaction (PCR); pre-analytics.

*Corresponding author: Sylvain Lehmann, CHU Montpellier, IRB, 80 Avenue Augustin Fliche, Montpellier 34295, France, E-mail: [email protected]

Sylvain Lehmann, Constance Delaby, Jérôme Vialaret and Christophe Hirtz: CHU Montpellier, Institut de Recherche en Biothérapie, Hôpital St Eloi, Laboratoire de Biochimie Protéomique Clinique et CCBHM, Montpellier, France; Université Montpellier 1, Montpellier, France; and INSERM U1040, Montpellier, France Constance Delaby: Université Paris 7-Denis Diderot, Paris, France Jacques Ducos: CHU Montpellier, Unité de Virologie Lapeyronie, Montpellier, France; and INSERM U1058, Montpellier, France

Introduction

Over a century since a new blood sampling method based on the use of a dry matrix was first described by Ivar Bang [1], the interest in dried blood spot technology has continuously evolved. This alternative approach, based on collecting blood spots on blotting paper and drying them, is called “dried blood spot” or DBS. In 1963, Robert Guthrie used this technique to develop systematic neona- tal screening for the metabolic disease, phenylketonuria [2]. Set up for the first time in Scotland, this use of DBS

spread to the UK in the 1970s, mainly to detect any innate errors in metabolism that were treatable. Of note, the use of DBS remains almost exclusively limited to this type of neonatal screening, even though many studies dem- onstrate its potential application in clinical biology, as well as in research. Indeed, classical clinical chemistry methods, small molecule and lipid analysis or molecular biology approaches, are all perfectly suited to the use of DBS. However, one limitation is represented by the small blood volumes associated with DBS sampling (5–10 µL) and therefore the need for very sensitive methods. Recent technological advances, in microfluidics, multiplex immunological/genomic detection systems, and mass spectrometry, could however settle most sensitivity prob- lems. In this overview we will summarize the pros and cons of this particular biological sampling method and evaluate its future role in clinical biology.

General DBS procedure

Collection and sampling

The collection area (finger, heel) has to be first disinfected.

The skin is then punctured with a sterile lancet (Figure 1).

The first blood drop is dabbed and subsequent drops are placed on blotting paper marked with circles to be filled.

Once all the required circles are filled, the blotting paper is left to dry for a few hours at room temperature on a non- absorbent surface. The drying time is very important as residual humidity favors bacterial development or molds and modifies the extraction stage [3].

Conservation

Once dry, the DBS cards are moved into a waterproof

plastic bag, possibly along with a desiccant and a humid-

ity indicator [4]. The purpose of the desiccant is to finalize

the drying process, which also minimizes any risk of infec- tion associated with sampling. Periods of storage at room temperature vary according to the biological factor, from 1 week for proteins [5], to 1 year or more for nucleic acids [6]. As far as serology is concerned, the blotting papers are usually kept at –20°C upon receipt [7]. For long-term preservation (up to several years) the blotting papers are stored either at −20°C or –80°C [8, 9].

Extraction

Extraction of the analytes from DBS specimens needs to be achieved using a standard procedure. One or more 2–8  mm diameter discs are then created with a specific punch. These small “spots” are placed in an elution buffer for variable time spans according to the procedure. The DBS extraction is then treated as a hemolyzed whole blood sample, and tested with methods often intended for plasma or serum. The elution buffer plays a major role in re-solubilizing the analytes to be tested. A wide variety of buffers are described in the literature. The most common are saline/phosphate buffers, often with added detergents (Tween, Triton…), carrier proteins and chelators [ethyl- ene diamine tetra acetic acid (EDTA)], as well as organic buffers with methanol, acetonitrile or ethanol. For nucleic acids, standard commercial kits exist which are compat- ible with molecular biology tests, from polymerase chain reaction (PCR) to genomic chips [10].

Patient Disinfection of

the sampling area

Prick with a lancet

Deposit on

filter paper Drying 1 to 3

hours at RT Transport/

Mailing

Punch (2-6 mm

diameter) Extraction with Analyses

appropriate buffer

Figure 1 DBS collection process.

Peripheral blood is collected by the patient at home. He disinfects the area (finger) and pierces the skin using a sterile lancet before blotting the blood onto high quality filter paper. The DBS is dried for 1–3 h at room temperature and mailed using the classical enve- lope. At the laboratory, the DBS is stored at room temperature. The sample is punched (2–6 mm) and the analytes are extracted using an appropriate buffer before analysis.

5x50µl 5

1500 rpm 5-10 ml

Dry Storage 4°C

y

Serum orPlasma

Storage Ambient T°

Whole blood with hemolysis Whole blood with

cell preservation VS.

Figure 2 Comparison of the use of classical blood sampling versus DBS sampling resulting in a 100-fold reduction in blood volume and an ease of storage.

Pros and cons of DBS

One of the main advantages of using DBS technology is that it allows access to samples in pre-analytical situations were standard blood collection is challenging (problem with sampling, storage). The typical DBS contains approx- imately 50 µL of whole blood on an average surface of 12 mm

(Figure 2). It enables the testing of various analytes such as nucleic acids, proteins, lipids, or small organic and non-organic molecules (Table 1). Two types of DBS are mostly available: cotton paper filters of different qualities (Whatmann 903 Protein Saver Cards Whatmann, Spring- field Mill, UK; Perkin Elmer 226 Spot Saver Card, Perkin Elmer, Waltham, USA) and glass microfiber filter papers (Agilent Bond Elut DMS, Santa Clara, CA, USA; Sartorius Glass Microfiber Filters, Goettingen, Germany). The main difference between the two supports is that the glass fiber does not soak up reagents, which diminishes non-specific analyte adsorption on the membrane.

In comparison to conventional blood testing, DBS

offers practical, clinical and financial advantages. Firstly,

DBS collection is easy to perform and relatively painless

(Figure 1). It can be carried out by the patient at home,

without the need for specialized structures such as

medical laboratories. This sampling procedure is far less

invasive than venipuncture, therefore is better suited for

patients requiring numerous blood tests, such as those

with damaged/altered veins, the elderly or infants. The

use of DBS also minimizes the volume of blood taken

from patients. It has been shown that drying the blood

spot on blotting paper damages the capsid of viruses [HIV,

Cytomegalovirus (CMV), hepatitis C virus (HCV), human

Table 1 Overview of DBS card usage in clinical chemistry other than its use for neonatal screening.

Methods Parameter Clinical interest References

Exogeneous nucleic acid Real-time PCR

Q PCR Human herpesvirus type 6 Differentiation active human

herpesvirus type 6 infection from inherited HHV-6

[11, 12]

RT-PCR Human hepatitis C Monitoring hepatitis C virus

(HCV) infection among injecting drug users

[7, 13]

Real-time PCR Human hepatitis B Hepatitis B virus (HBV) DNA

quantification [14]

Real-time PCR, Q-PCR Cytomegalovirus Diagnosis of human

congenital cytomegalovirus infection

[15, 16]

Nested PCR, RNA assays, RT-PCR HIV virus Detection of human

immunodeficiency virus [8, 13, 17]

Peptides/proteines

ELISA HIV virus Human immunodeficiency

virus serotyping [18]

ELISA C-reactive protein Cardiovascular risk [19]

DELFIA Free-β human chorionic gonadotrophin

(free-β hCG) and PAPP-A Fetal aneuploidy risk [20]

Immuno-fluorometric assays Luteinizing hormone and follicle-

stimulating hormone Circulating gonadotropin

concentrations [21]

Chemiluminescent

immunoassay Prostate specific antigen (PSA) Prostate cancer screening [22]

RIA Somatedin-C (IGF-1) Screening test for growth

hormone deficiency [23]

ELISA Apoliproteins B Hypercholesterolemia [24]

Immune nephelometry α

-Antitrypsin α

-Antitrypsin deficiency [5]

ELISA α-Fetoprotein Open neural tube defect and

Down syndrome [25]

Enzyme assays Biotinidase Biotinidase deficiency [26]

EIA Calcitonin gene-related peptide Children with autism or

mental retardation [27]

LC-MS/MS Ceruloplasmin Wilson’s disease [28]

Spectrophotometry Hemoglobin Folate analysis [29]

Turbidimetric immunoassay Glycated hemoglobin A1c Diagnosis and treatment of

diabetes [30]

LC-MS/MS HbA

Diagnosis of thalassemia [31]

Non-radiochemical HPLC Hypoxanthine-guanine

phosphoribosyltransferase adenine phosphoribosyltransferase adenosine deaminase

Purine metabolism disorders [32]

LC-MS/MS Iduronate 2-sulfatase Diagnosis of Hunter disease [33]

ELISA, RIA Insulin-like growth factor Evaluation of growth hormone

status [34]

ELISA Prolactin Diagnosis of epilepsy [35]

ELISA Transferrin receptor Iron deficiency [36]

DELFIA Thyroglobulin Thyroid status [37]

ELISA CD4 CD4+ lymphocyte counts in

HIV patients [38]

ELISA Measles and rubella IgM and IgG Detection of measles and

rubella IgM and IgG [39]

DELFIA Toxoplasma gondii-specific IgM and

IgA Screening of congenital

toxoplasmosis [40]

RIA Insulin Diagnosis of hyperglycemia/

hyper-insulinemia [41]

Enzyme assays Acid α-glucosidase Glycogen storage disease II [42]

Methods Parameter Clinical interest References

Enzyme assays 8 lysosomal enzymes Clinical differentiation among

mucopolysaccharidosis, oligosaccharidosis, and mucolipidosis II and III

[43]

Enzyme assays α-iduronidase activity Diagnosis of α-L-iduronidase

deficiency [44]

Biochemistry Phytanic acid and pristanic acid Diagnosis of peroxisomal

disorders [45]

Electro-immunodiffusion β-Lipoprotein Familial type II and combined

hyperlipidemia [46]

ELISA Fumarylacetoacetase Hereditary tyrosinemia type I [47]

Luminex TGF-β1, (MCP-1, (MIP-1α, MIP-1β,

NT-4, BDNF, RANTES, CRP, MMP-9… Inflammatory status [48]

Enzyme immunoassay IgE Allergic disease and repeated

macro-parasitic infections [49]

ELISA IgG and IgA Nasopharyngeal carcinoma

screening [50]

Enzyme assays Lysosomal b-d-galactosidase (bG; EC

3.2.1.23) Mucopolisaccharidosis type I [51]

Fluorometric immunoassay Thyroid-stimulating hormone Immunoreactive trypsin, creatine kinase MM isoenzyme

Congenital hypothyroidism, congenital adrenal hyperplasia, and muscular dystrophy

[52]

Column chromatography Thyroxine-binding globulin Neonatal hypothyroidism [53]

Immunoassay Trypsine immunoreactive (IRT) Cystic fibrosis [54]

ELISA Antibodies against hepatitis A Hepatitis A [55, 56]

ELISA Antibodies against hepatitis B Hepatitis B [57]

CORECELL Maternal antibody to hepatitis B Infection with HBV [58]

ELISA Anti-HCV antibodies Detection of antibodies to

hepatitis C virus [59, 60]

ELISA Anti-malarial antibodies Diagnosis of malaria [61]

ELISA

patients with cystic fibrosis [62]

ELISA Thyroid antibody Thyroid-antibody screening [63]

ELISA Antibodies against tetanus Screening of tetanus and

diphtheria toxins [64]

ELISA Antibodies against Brucella Diagnosis of human

brucellosis [65]

ELISA Antibodies against cysticercus Detection of anti-cysticercus

antibodies [66]

ELISA Antibody against HTLV-1 and HTLV-2 Detection of the Human

T-lymphotropic virus [67]

Immuno-fluorescence Antibodies against to Coxiella burnetii,

Diagnosis of Rickettsial

diseases [68]

ELISA Antibody against syphilis Diagnosis of syphilis [69]

Indirect hemagglutination test Antibody against Treponema Diagnosis of syphilis [70]

ELISA Antibody against Trypanosoma cruzi Diagnosis Trypanosoma cruzi

infections [71]

ELISA Antibody against Trichomonas

vaginalis

Seroepidemiology of

Trichomonas vaginalis

[72]

Fluorescent Galactose-1-phosphate

uridyltransferase (GALT) Galactosemia [73]

ELISA Epstein-Barr virus Epstein-Barr virus

immunoglobulin G (IgG) serology

[50]

EIA Rubella virus Detection of congenital

Rubella virus [74]

(Table 1 Continued)

Methods Parameter Clinical interest References

EIA Dengue virus Dengue virus diagnosis [75]

ELISA Antibodies against hepatitis A Hepatitis A [55, 56]

ELISA Antibodies against hepatitis B Hepatitis B [57]

CORECELL Maternal antibody to hepatitis B Infection with HBV [58]

ELISA Anti-HCV antibodies Detection of antibodies to

hepatitis C virus [59, 60]

Multiplex ligation-dependent probe amplification on DNA (MLPA)

Detecting 22q11.2 deletions Manifestations associated

with DiGeorge syndrome [76]

PCR GSTM1 et GSTT1 gene variant Researching pediatric cancer

susceptibility genes [77]

ELISA multiplex Human papillomaviruses (HPV),

Helicobacter pylori, hepatitis C virus

(HCV), and JC polyomavirus (JCV)

Infections of HPV, H. pylori,

HCV, and JCV [78]

Lipids and small molecules

Densitometry Phenylalanine Phenylketonuria [2]

Enzymatic method Triglycerides Evaluation of the

cardiometabolic risk [79]

LC-MS/MS Amino, organic, and fatty acid Metabolic disorders [80]

Fluorimetric HPLC method Homocysteine Homocysteinuria [81]

Enzymic methods Determination of glucose Monitoring of diabetic

patients [82]

LC-MS/MS 17-OHP, androstenedione Congenital adrenal

hyperplasia [83]

HPLC Retinol Retinol analysis [84]

LC-MS/MS Thyroxin (T4) and TSH Congenital hypothyroidism [85]

Chemiluminescence Free thyroxine (FT4) Assessment of thyroid status [86]

LC-MS/MS Free carnitine Inborn errors of metabolism [87]

GC-MS Methylcitrate Newborn screening for

propionic acidemia [88]

GC-MS Octanoate, decanoate, cis-4-decenoic

acid (C10:1) and cis-5-tetradecenoic acid

Free fatty acids [89]

LC-MS/MS Succinylacetone Hepatorenal tyrosinemia [90]

FIA-ESI-MS/MS Guanidinoacetate and creatine Primary creatine disorders [91]

Xenobiotics

LC-MS HIV antiretroviral drugs

(NVP, SQV, ATV, APV, DRV, RTV, LPV, EFV, ETV)

HIV therapeutic follow-up [92, 93]

RIA Cocaine metabolite (benzoylecgonine) Information on newborns and

maternal exposures to various substances, including drugs of abuse

[94]

LC/MS Quinine, mefloquine, sulfadoxine,

pyrimethamine, lumefantrine, chloroquine

Blood levels of drugs administered for malaria and pneumonia treatment

[95, 96]

Capillary gas chromatography Dichlorodiphenyldichloroethylene Newborns’ body burden of

environmental pollutants [97]

Fluorescence polarization

immunoassay Theophylline Therapeutic drug monitoring [98]

Genomics

PCR Mutations of factor V G1691A (FVL),

prothrombin (PT) G20210A, 5′10′

methylenetetrahydrofolate reductase (MTHFR) C677T, and methionine synthase (MS) A2756G

Susceptibility to venous

thromboembolism [99]

Real-time PCR Mutation c.-32T > G (IVS1-13 > G) Acid maltase deficiency [100]

DNA-based assay Mutation (IVS4+919G- > A) Fabry disease [101]

(Table 1 Continued)

T-lymphotropic virus (HTLV)] [108] reducing any possible risk of contamination for medical or paramedical staff [4].

In addition, it enables the shipping of samples by regular mail with no particular risk of contamination. This repre- sents a valuable asset for sampling in remote communi- ties either located far away from a testing laboratory or with limited technical infrastructure available, therefore provides added value compared to standard blood sam- pling [59]. Through its small size and stacking capacity, DBS is also a valuable solution for reducing and facilitat- ing storage in clinical laboratories and biobanks [109]. It is noteworthy that in case of storage, an individual bagging or a separation using a sturdy paper will be important to avoid the possibility of cross-contamination between cards [3]. These properties of DBS have been utilized in experimental research, by facilitating pharmacological studies and pharmacokinetics on small animals with very limited volumes of biological liquids. This follows the reg- ulations aimed at protecting small animals (decreasing sample volume and sophistication of sampling methods) during pre-clinical studies [110]. Concerning sample sta- bility, many studies have shown that most analytes from whole blood are stable at room temperature for at least 7 days. In some cases such as opiates, DBS even increases stability during storage [111], and nucleic acids are a major tool for short- and long-term preservation, as they can be isolated after several months at room temperature and several years at −20°C. [112]. From a medico-economi- cal point of view, it is interesting to note that the use of DBS allows a significant cost reduction due to decreased requirements in trained staff, facilitated transportation, storage, and processing.

A major drawback of DBS technology resides in the nature of the biological sample itself (Figure 2). In a stand- ard sampling procedure, either serum or plasma is ana- lyzed, whereas DBS samples are composed of hemolyzed whole blood. Hence, interferences due to hemoglobin and the release of intracellular content could occur. The blood cells (erythrocytes, leukocytes, platelets etc.) are altered by the drying process, thus cellular hematological testing is impossible. Drying can also denature proteins and alters the enzymatic activity of blood proteins (aspartate transaminase). Any remaining cells in the samples can also change the global protein composition and therefore modify the concentration of some analytes. In some cases, clinical thresholds set up using standard blood samples need to be adapted. Hematocrit that affects blood dis- persal on the blotting paper also needs to be taken into account [113]. The small volume of samples resulting from the DBS can be a disadvantage for low sensitivity assays [4] and for running multiple tests.

Use of DBS in clinical chemistry

The primary use of DBS in France is systematic neona- tal screening. As blood sampling in newborns is diffi- cult, DBS technology represents a viable alternative. DBS testing was set up in 1978 by the French Association for screening and preventing disabilities in children (http://

www.afdphe.org/). Sampling of newborns enables the detection of phenylketonuria, hypothyroidism, adrenal hyperplasia, cystic fibrosis and sickle cell disease (in

Methods Parameter Clinical interest References

DHPLC Substitution (c.840C > T) Spinal muscular dystrophy [102]

Specific restriction digest

method Mutation (c.985A > G) Medium chain acyl-coA

dehydrogenase deficiency (MCADD)

[103]

PCR Mutation of cystic fibrosis

transmembrane conductance regulator (CFTR)

Cystic fibrosis [104]

PCR DNA mutation β-thalassemia [105]

PCR

Real-time PCR SMN1 exon 7 deletions

Copy number variations of SMN1 and SMN2

Spinal muscular atrophy [106]

PCR FMR1 methylation Fragile X syndrome [107]

Multiplex ligation-dependent probe amplification on DNA (MLPA)

Detecting 22q11.2 deletions Manifestations associated

with DiGeorge syndrome [76]

PCR GSTM1 and GSTT1 gene variant Researching pediatric cancer

susceptibility genes [77]

(Table 1 Continued)

some areas). The extension of these tests to cover a wider number of diseases, similar as in to USA, is currently under consideration [28]. A positive result will always be confirmed or denied by further specific tests. Beyond its use for neonatal screening, many clinical analytes can be measured using DBS. These analytes are divided into four major categories as follows (see also Table 1).

Exogenous nucleic acids

The measurement of nucleic acids is typically required in the virology field. There is a growing interest in viral screening through nucleic acid detection (RNA, DNA) using DBS, as current molecular biology technologies [quantitative polymerase chain reaction (Q-PCR), reverse transcription polymerase chain reaction (RT-PCR)] are very sensitive and require only a small sample amount ( < 20 µL). Nevertheless, it is important to note that the amount of material available from a DBS sample is between 1 and 2 logs lower compared to a standard serum or plasma sample. The preservation of nucleic acids on blotting paper is stable for long periods [3], providing it is dried and stored away from humidity in a suitable plastic bag containing a desiccant. DBS nucleic acid detection is mainly used in screening for viral diseases such as cyto- megalovirus [15], herpes simplex virus [11], hepatitis A [55], hepatitis C [13] and HIV [114].

Peptides – proteins

Concerning proteins and peptides one caveat is repre- sented by the relative difficulty of their extraction from DBS samples, as well as the low sensitivity of certain protein dosage. The main proteins measured from DBS can be classified into two groups: standard serum pro- teins and antibodies. The most widely used analytical techniques are immunological assays which measure clinical analytes with good specificities and sensitivities.

An example is represented by the immunoturbidimet- ric assay for glycated hemoglobin (to monitor glycemic balance in diabetic patients). Glycated hemoglobin meas- ured from DBS samples correlate well with standard tests.

In addition, this analyte remains stable for over 15 days on DBS [30]. DBS is also well adapted for the enzyme-linked immunosorbent assay (ELISA) detection of specific anti- bodies against Epstein-Barr virus [50], Rubella virus [74], dengue virus [75] or hepatitis C [7, 59] and HIV virus [13].

An interesting evolution of liquid chromatography/

mass spectrometry (LC/MS) is represented by quantitative

techniques for measuring peptides and proteins [115]. This approach was adapted on DBS to measure ceruloplasmin for the neonatal screening of Wilson’s disease [28] and for peptide C quantification [116]. When used in multiplex mode (multiple reaction monitoring) this mass spectrom- etry method has the potential to measure many analytes within only a few microliters [115]. For instance, Chambers et al. [117] have succeeded in quantifying a panel of 40 serum proteins from DBS, using this approach.

Lipids, sugars and small molecules

Phenylalanine, an amino acid measured in phenylke- tonuria screening of newborns, exemplifies the dosage of small molecules using DBS [2]. Small organic mol- ecules are significantly less sensitive than proteins to the drying process which characterizes DBS samples. In addition, the major progress of liquid chromatography/

mass spectrometry (LC/MS) in this field has allowed the quantification of many small molecules such as vitamin D [118] or lipids [79]. For instance, high levels of triglyc- erides, representing an important risk for cardiovascu- lar and coronary diseases, can be quantified using DBS.

These analytes remain stable on DBS for 30 days at room temperature and up to 90 days at 4°C. The profiling of glycans on DBS was also recently achieved using mass spectrometry [119].

Xenobiotics

In 1993, Henderson et al. [120] demonstrated the use of

DBS for detecting narcotics, such as cocaine, through

modification of a radioimmunoassay (RIA) commercial

kit. Xenobiotic testing using DBS has since played an

important role, mainly by the screening of antimalarial

and antiretroviral drugs by LC/MS in isolated popula-

tions [95]. Another example is represented by the quan-

tification of nine antiretroviral molecules in HIV using

DBS. This detection method has been validated by the

Food and Drug Administration (FDA) with sample stabil-

ity ranging from 12 to over 90 days at room temperature

[92]. In the field of toxicology, which is a major applica-

tion of DBS [121], Saussereau et al. [122] have, for example,

developed a new drug screening method based on LC/MS

using on-line extraction for the quantification of opiates,

cocainics or amphetamines. The development of these

new measurement techniques, based on LC/MS for xeno-

biotics, will greatly increase the interest of using DBS in

clinical chemistry.

Genomics

The clinical potential of DBS for genomics has been dem- onstrated as early as 1987 [123]. DNA micro-extraction from dried blood has allowed the detection of mutations responsible for diseases such as cystic fibrosis [124], X fragile syndrome [107], spinal muscular atrophy [106], cancers [77] and thalassemia [105]. DBS, which is a fairly inexpensive sampling and storage method, is also a good choice for genetic material biobanks [125]. For instance, the Danish national biobank for neonatal screening includes over 2 million DBS which virtually corresponds to all Danish people born since 1982.

Conclusions

The use of DBS has many advantages in terms of sampling, transportation, storage and biosafety when compared to classical collection methods. One interesting aspect of DBS is the possibility of simplified “self/home blood sampling”. The patient will be able to independently and safely collect a blood sample. The DBS will then be sent to the laboratory by mail. As described in this review, many clinical analytes are already available on DBS, and more are to follow, thanks to innovative approaches. Indeed, development of microfluidics, multiplex immunological/

genomic detection systems, mass spectrometry and automated DBS processing open new interesting clini- cal prospects. The detection and follow-up of metabolic, infectious and chronic diseases could therefore rely on the use of DBS. Both the patient and society could benefit from this technology. Already, several public and com- mercial laboratories in both Europe and USA are offer- ing DBS kits for a broad range of analytes often grouped into panels for hormonal, metabolic or cardiovascular diseases. This evolution could dramatically change how clinical chemistry pre-analytics are handled in the future.

Acknowledgments: The authors thank Rachel Almeras, Bader Al Taweel, Domitille Héron and Thibault Fortane for their initial help in the writing of this review and Brigitte Lehmann for editing the manuscript.

Conflict of interest statement

Authors’ conflict of interest disclosure: The authors stated that there are no conflicts of interest regarding the publication of this article.

Research funding: None declared.

Employment or leadership: None declared.

Honorarium: None declared.

Received March 26, 2013; accepted April 19, 2013; previously published online June 1, 2013

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Constance Dalaby holds a PhD in Biology and graduated from University Paris 7 (France) in 2006. In 2005, she was recruited as a research assistant in Biochemistry by the Clinical Unit of Biochemistry at Pr Jean-Charles Deybach (Hôpital Louis Mourier, Colombes, France), as a member of the French Reference Center of Porphyrias. In 2008–2009, she collaborated with the Biochemistry Unit of the Hospital Clinic of Barcelona (Dr Jordi To-Figueras, Spain) and was recruited as an Assistant Professor in Biochemistry and Molecular Biology by the University Paris 7 (France) in 2009.

Since 2011, she has been part of the Clinical Chemistry Laboratory (Proteomics Platform) of Pr Sylvain Lehmann (IRB, Hôpital Saint Eloi, CHRU Montpellier).

Prof. Sylvain Lehmann trained as an MD/PhD (1992, Strasbourg,

France). He was the recipient of a Howard Hughes fellowship for

physicians and spent 4 years in Washington University, St Louis,

MO, USA, as a postdoctoral fellow and a research Assistant Profes-

sor. From 1997, he was a researcher of the French National Research

Institut (INSERM) and in 2003 he obtained a position as Professor

of Biochemistry at the Medical School of Montpellier (France). His

research focuses on neurodegenerative disorders (Alzheimer, prion,

etc.) and on clinical proteomics. He is a vice-president of the French

National Society of Clinical Biology (SFBC) in charge of its Scientific

Committee and he chairs the Working Group “Clinical Quantitative

Mass Spectrometry Proteomics” of the International Federation of

Clinical Chemistry and Laboratory Medicine (IFCC).

Christophe Hirtz was born in 1972 and studied Biochemistry at Paul Sabatier University in Toulouse. Having got his Doctorate and an accreditation to supervise research in the proteomic field, he was recruited in 2008 as an Associate Professor at the University Mont- pellier I and is specialized in Biochemistry and Clinical Proteomic.

His scientific interest includes the development of a new method of protein quantification using targeted mass spectrometry in a clini- cal environment. He is in charge of the Clinical Proteomic Platform of the Laboratory of Biochemistry and Clinical Proteomic directed by Pr. Sylvain Lehmann in Montpellier.

Jérôme Vialaret was born in 1984 and studied Organic Chemistry at Montpellier II University (obtaining his Master’s degree in 2007). He specialized in proteomic while working for Pierre Fabre Laboratories, EPFL (Lausanne, Switzerland), INRA (Montpellier, France) and at Montpellier Hospital. Having amassed a wealth of experience in large scale proteomic (proteome and phosphopro- teome) with dedicated quantitative methods (silac and label-free), he focused on the development of protein quantification using targeted mass spectrometry in a clinical environment. He is in charge of these method developments in the Clinical Proteomic Platform of the Laboratory of Biochemistry and Clinical Proteomic directed by Pr. Sylvain Lehmann in Montpellier.

Dr. Jacques Ducos was trained as an MD (1983, Montpellier, France) and PhD (1993, Montpellier, France). He was a resident in the Montpellier Hospital (1979–1983). He specializes in Virology (HBV/

HCV/HIV) and his research is focusing on biological markers of viral

infections. He is currently responsible for the functional unit of viral

hepatitis at the Montpellier Hospital (Lapeyronie, France). He is the

president of the Viral Hepatitis Network of the Languedoc Roussillon

Camargue (RHEVIR).