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Use of Synthetic Peptides and Multiple Antigen Blot Assay in the

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VIM-2018-0023-ver9-Gauna_1P
Type: research-article
VIRAL IMMUNOLOGY
Volume 31, Number 8, 2018
ª Mary Ann Liebert, Inc.
Pp. 1–7
DOI: 10.1089/vim.2018.0023

Use of Synthetic Peptides and Multiple Antigen Blot Assay

in the Immunodiagnosis of Hepatitis C Virus Infection

AU1 c Adriana Gauna,1 Sandra Losada,2 Maria Lorenzo,2 Marilyan Toledo,3 Henry Bermúdez,2
Pierina D’Angelo,4 Doneyla Sánchez,4 and Oscar Noya2,5

Abstract

Acute hepatitis C virus (HCV) infection is usually asymptomatic, therefore, early diagnosis is rare. It may
remain undiagnosed in individuals who progress to chronic infection, often until serious liver damage has
developed. To incorporate the diagnosis of this viral disease in a multiple-diagnostic assay, we first analyzed by
immunoinformatics the HCV subtype 1a polyprotein (specifically Core, E2, NS3, NS5A proteins) to select
antigenic peptides to be tested initially by the Pepscan technique. Next, we performed the immunodiagnosis of
HCV infection, using the Multiple Antigen Blot Assay (MABA). In 22 patients’ sera included in this study, a
20-mer linear peptide belonging to the N-terminus of the worldwide conserved Core protein showed 100%
sensitivity and specificity; other sequences showed different levels of antibody recognition. The use of MABA
in combination with synthetic peptides as a source of multiple, specific, and nonexpensive antigens for other
infectious diseases could represent a rapid, integrated, and inexpensive diagnostic methodology.

Keywords: hepatitis C virus, synthetic peptides, immunodiagnosis, MABA

Introduction Initial cleavages release the individual envelope proteins (E1

and E2) and the capsid constituent protein (Core or C), located in

S ince its discovery in 1988 (6), the Hepatitis C Virus

(HCV) has become a target of intense research, which is
not surprising given the current statistics. Around the world,
there are more than 150 million infected people and between
300,000 and 500,000 annual deaths due to severe hepatic
disorders related to this infection (28).
HCV is an enveloped virus, composed by a positive sense
RNA strand, classified in the Flaviviridae family (18). The en-
velope consists of two proteins (E1 and E2) that surround the
nucleocapsid, which is composed of multiple copies of a small
basic protein called core, containing the genomic RNA (16).
After this genomic RNA enters the cytoplasm of the hepato-
cytes, the translation process occurs immediately, and the re-
sulting polyprotein has 3,010–3,033 amino acids, depending on
the virus genotype (7,15). This polyprotein is subsequently
cleaved, yielding structural and nonstructural viral proteins (18).
the N-terminal region of the polyprotein. The remaining seg-
ment, containing viral enzymes or nonstructural proteins (NS2,
NS3, NS4A, NS4B, NS5A, and NS5B), is released by viral
proteases in subsequent cleaving events (12,13,29).
Effective evaluation and rapid and accurate diagnosis of
HCV infection are extremely relevant in the control of the
transmission and opportune treatment of infected patients.
Exposure to the virus can be determined with high sensi-
tivity and specificity with the currently available indirect
serological assays that include multiple recombinant antigens
from Core, NS3, NS4, and NS5 proteins (1,17). However,
the diagnosis of HCV infection remains a subject of study
because these assays are expensive, especially for developing
countries. In this sense, searching for antigenic peptides de-
rived from proteins traditionally used in current diagnosis kits
can provide a less expensive alternative.

1
Programa de Doctorado en Biotecnologı́a, Pontificia Universidad Católica de Valparaı́so/Universidad Técnica Federico Santa Marı́a,
Valparaı́so, Chile.
2
Sección de Biohelmintiasis, Instituto de Medicina Tropical, Facultad de Medicina, Universidad Central de Venezuela, Caracas,
Venezuela.
3
Cátedra de Parasitologı́a, Escuela de Medicina ‘‘Luis Razetti,’’ Universidad Central de Venezuela, Caracas, Venezuela.
4
Laboratorio de Programas Especiales-Hepatitis y SIDA, Dpto de Virologı́a, Gerencia Sectorial de Diagnóstico y Vigilancia
Epidemiológica, Instituto Nacional de Higiene ‘‘Rafael Rangel,’’ Caracas, Venezuela..
5
Centro para Estudios Sobre Malaria, Instituto de Altos Estudios ‘‘Dr. Arnoldo Gabaldón’’ Instituto Nacional de Higiene-Ministerio del
Poder Popular para la Salud, Caracas, Venezuela.

1
VIM-2018-0023-ver9-Gauna_1P.3d 08/06/18 9:16am Page 2

2 GAUNA ET AL.

To select candidate diagnostic peptide epitopes, we ana-
lyzed the sequence of 2 structural (Core and E2) and 2
nonstructural (NS3 and NS5) proteins of 22 selected pep-
tides that include several mutations found in the GenBank
Database. We combined two sequential strategies known as
the Pepscan (27) and MABA (Multiple Antigen Blot Assay)
(21,22) methodologies, which have been successful in the
identification of an antigenic sequence for hepatitis A virus
(HAV) diagnosis (10). The first strategy consists of the
synthesis of a library of peptides (possible antigens) in a
Spot manner attached to a cellulose membrane support to
subsequently be confronted with patients’ sera. The most
antigenic peptides were selected, subsequently synthesized
in greater quantity and then explored by MABA.
Materials and Methods
Patient samples and ethical clearance
Twenty-two characterized sera with informed-consent
signature were provided by the Laboratorio de Hepatitis y
SIDA of the Departamento de Virologı́a of the Instituto
Nacional de Higiene ‘‘Rafael Rangel’’ (Ministerio del Poder
Popular para la Salud-Venezuela). The diagnosis of hepatitis
C patients was accomplished by serological third-generation
tests, and those borderline cases were referred to INNO LIA
HCV, an enzyme immunoassay for verification of anti-
bodies against specific proteins of the virus, and finally by
polymerase chain reaction of the 5’NC. This study followed
both the international and national standards on human ex-
AU2 c perimentation, including the Declaration of Helsinki (WMA
2000) and national regulations. This project has been eval-
uated and accepted by the Bioethics Committee of The In-
stituto de Medicina Tropical (IMT) of the Universidad
Central de Venezuela.
Protein selection and prediction of B-cell epitopes
The HCV polyprotein subtype 1a (GenBank accession
No. AGN33396.1), specifically Core, E2, NS3, and NS5A
proteins, was selected for this immunoinformatic analysis.
For the determination of possible linear B-cell epitopes,
ANTHEPROT (8) was used, based on the physicochemical
characteristics of amino acids, such as hydrophobicity (14),
surface accessibility (3), and antigenicity (23). In the case
of the NS3 protein, a crystallized structure (PBD: 1CU1) is
available (30), which was used to locate the peptides se-
lected by ANTHEPROT to adjust the epitopes. Crystal-
lized structures are also reported for the NS5A protein
[pdb: 1ZH1 (26) and 3FQM (19)]. Furthermore, peptides
were additionally analyzed by multiple alignment with
other polyprotein sequences (GenBank accession No.
ABV46185.2, AGN33294.1, AEW90302.1, ACJ37189.1)
using Muscle tool (www.ebi.ac.uk/Tools/msa/muscle) (24)
and some mutations were included, represented as bold and
underlined residues in Table 1. b T1
Peptide synthesis on cellulose paper
Twenty-two peptides were manually synthesized using
the Spot technique as described previously (10,27) on a
Whatman 50 cellulose paper. After the peptide sequences
were assembled, the side-chain protecting groups were
removed with two cleavage solutions for 30 min and 3 h,
respectively, without agitation. Solution I consists of 90%
trifluoroacetic acid (TFA), 5% ultrapure water, 3% triiso-
propylsilane (TIPS) and 1% phenol and 1% dichloromethane
(DCM). Solution II consists of 50% TFA, 2% ultrapure
water, 3% TIPS, 1% phenol, and 44% DCM. The re-
agents were from Iris Biotech and Sigma-Aldrich che-
mical grade.

Table 1. Sequences Selected for Spot Synthesis and Selection Criteria

Spot (peptide) Protein Sequence Position Size (mer) Selection criteria
1 Core MSTNPKPQRKTKRNTNRRPQ 1–20 20 PF
2 RLGVRTTRKTSERSQPRGRR 43–62 20 PF
3 SQPRGRRQPIPKARRPEGRT 57–75 20 PF
4 PSWGPTDPRRRSRNLGKVID 105–123 20 PF
5 E2 VPTYNWGENETDVFVLNNTR 524–543 20 PF
6 VPTYSWGENETDVFVLNNTR 524–543 20 PF/mutation
7 NWTRGERCDLEDRDRSELSP 645–664 20 PF
8 NS3 ITSLTGRDKNQVEGEVQIVS 1044–1063 20 PF
9 DVIPVRRRGDSRGSLLSPRP 1138–1157 20 PF
10 GRHLIFCHSKKKCDELAAKL 1388–1407 20 PF
11 APHGARSLTPCTCGSS 1112–1128 16 TS
12 APPGARSLTPCTCGSS 1112–1128 16 TS/mutation
13 SPVFTDNSSPPAVPQSF 1207–1223 17 TS
14 LHGPTPLLYRLGAVQNEVT 1618–1635 19 TS
15 NS5A SHITAEAAGRRLARGSPPSV 2178–2198 20 PF
16 SHITAEAAGRRLARGSPPSL 2178–2198 20 PF/mutation
17 ITRVESENKVVILDSFDPLV 2241–2260 20 PF
18 ITRVESENKVVVLDSFDPLV 2241–2260 20 PF/mutation
19 AEEDEREISVPAEILRKSRR 2261–2280 20 PF
20 PVPPPRKKRTVVLTESTVST 2322–2342 20 PF
21 GSPPSVASSSASQLSAPSLK 2192–2212 20 PF
22 GSPPSLASSSASQLSAPSLK 2192–2212 20 PF/mutation
Bold and underlined residues correspond to mutations included by multiple alignment analysis.
PF, physicochemical profiles; TS, tertiary structure.
VIM-2018-0023-ver9-Gauna_1P.3d 08/06/18 9:16am Page 3
USE OF SYNTHETIC PEPTIDES AND MABA IN HCV IMMUNODIAGNOSIS
Immunoassay with cellulose-bound
peptides (Pepscan)
The dry paper with the ‘‘spots’’ of peptides was blocked
with phosphate-buffered saline containing 0.05% Tween-20
(PBST) and 5% nonfat dry milk. Peptides were tested with a
pool of four hepatitis C patients’ sera (diluted 1:250 each).
Horseradish peroxidase (HRP)-conjugated anti-human IgG
(Santa Cruz Biotechnology) was used as the secondary an-
tibody and developed with a solution of Luminol Super-
Signal West Pico substrate (Thermo Scientific). The output
chemiluminescent signal was recorded using ChemiDoc
Bio-Rad equipment.
Solid-phase peptide synthesis
Based on the results obtained with the cellulose-bound
peptides, those that showed antibody recognition were syn-
thesized by a manual solid-phase Fmoc strategy in polypro-
pylene syringes fitted with a polyethylene porous disk (2,4,9).
The first amino acid was coupled manually as follows: 2-
chlorotrityl resin (0.4 mmol, 1.0 mmol/g) was washed with
DCM (3 · 1 min) and swelled in DCM for 15 min (2 · 15 min).
Fmoc-aa-OH (1 equiv) and N,N-diisopropylethylamine (DI-
PEA) (3 equiv) were sequentially added to the resin, and the
mixture was allowed to react for 90 min. Next, a capping step
with methanol (MeOH)-DCM-DMF-DIPEA (5:40:40:15;
2 mL) was carried out. The rest of the synthesis was carried out
using 2-(1H-Benzotriazole-1-yl)-1,1,3,3-tetramethylaminium
tetrafluoroborate (TBTU) as a coupling reagent and DIPEA
as a base. Fmoc removal was performed with 20% 4-
methylpiperidine-DMF (1:4; 1 · 5 min, 1 · 10 min). Washings
between deprotection and coupling were carried out with
DMF (3 · 0.5 min) and DCM (3 · 0.5 min), using 2 mL of each
solvent.
Peptides were synthesized with and without incorporation
of stearic acid at the N-terminal end of the sequence. The
fatty acid coupling was carried out using 15 M excess of this
acid activated with N,N¢-Diisopropylcarbodiimide (DIC)/
hydroxybenzotriazole (HOBt) in DCM. Then, the complete
peptides were cleaved from the support by using TFA
(95%)/TIPS (2.5%)/ultrapure water (2.5%) for 3 h. Char-
acterization and purity were evaluated by Electrospray Io-
nization Time-of-Flight (ESI-TOF) mass spectrometry and
high-performance liquid chromatography, respectively.
Analysis of peptides antigenicity by the MABA
The simultaneous antigenicity of the synthetic peptides
was analyzed by a special dot blot assay developed in our
laboratory known as MABA (21,22). A rectangle of nitro-
cellulose (NC) paper (Bio-Rad) was cut and soaked in dis-
tilled water. The wet paper was located on an acrylic
template (Miniblotter 20SL) for its sensitization by the
introduction of 60 mL of different peptide preparations
(20 mg/mL in pH 9.6 carbonate-bicarbonate buffer) in each
parallel groove. In addition, during immobilization on the
membrane, a normal control human serum (HS) was in-
cluded in two of the parallel marginal lanes as an internal
control of the reaction in each strip. This control should
always react in the presence of the secondary antibody. The
peptide solutions were removed by washing, and blocking
was achieved by immersing the NC in 5% nonfat milk in
3
PBST. Once the NC was blocked, 2 mm-width-strips were
cut perpendicular to the channels, and each strip thus con-
tained a row of square spots corresponding to the different
antigens. Strips were immersed individually in the troughs
of an incubation tray in the serum (diluted 1:100 in blocking
solution) of a larger sample of hepatitis C patients’ sera
(including the four ones previously used in Pepscan). In this
preliminary study, we had access to sera from 22 hepatitis C
patients. Four negative sera and one no-serum control (No-
SC) were included. The reaction was detected separately with
two secondary antibodies. The first one consists of goat anti-
human IgG peroxidase (1:15,000; Santa Cruz Biotechnol-
ogy), which was developed using the Luminol SuperSignal
(West Pico Chemiluminescent Substrate) and recorded in a
ChemiDoc Bio-Rad equipment. The second one consists of
anti-human IgG alkaline phosphatase (1:15,000; Sigma) us-
ing a combination of nitro blue tetrazolium chloride (NBT)
and 5-bromo-4-chloro-3¢-indoly phosphate p-toluidine salt
(BCIP) as chromogenic substrate.
Results
Peptide selection
To select sequences with antigenic potential, four proteins
were analyzed by ANTHEPROT software and peptides of
20-mer with high values of hydrophilicity, solvent accessi-
bility, and antigenicity were chosen. Fifteen peptides were
selected based on their physicochemical profiles; four of
them belong to the Core protein, two to E2, four to NS3, and
five to NS5A. The E2 protein showed a less hydrophilic and
antigenic sequence. However, 20-mer peptides with the
higher possible values were selected. Figure 1 represents an b F1
example of sequence analysis, together with the ANTHE-
PROT profile of the Core protein and the selected peptides.
The X-ray structure of the NS3 protein is currently available
(PDB: 1CU1), which allowed us to determine if sequences
selected by primary structure predictions by ANTHEPROT
are exposed in the tertiary structure. This analysis showed
that all selected peptides are well structured and the two
sequences selected in the protease domains are exposed.
However, one of the peptides of the helicase domain was
discarded, as it was not exposed to solvents.
The reported crystallized structures of the NS5A protein
were not considered in this study. The possible epitopes
obtained by the ANTHEPROT analysis cannot be located in
the three-dimensional (3D) structures as they are completely
different sequences. The sequence used in this study is of
recent date (2014), while the 3D structures were reported in
2005 and 2009. We propose to use them in the future to
predict possible B epitopes based on structure.
Finally, even when the polyprotein of genotype 1a of
HCV is conserved, there are several available sequences,
which is the reason why a multiple alignment was utilized to
include some possible mutations. All 22 selected sequences
are summarized in Table 1, as well as their selection criteria.
Solid phase peptide synthesis and testing on cellulose
paper (Pepscan)
To experimentally determine the antigenicity of selected
peptides, the 22 sequences in Table 1 were synthesized
on a functionalized cellulose paper in a spot manner. Three
VIM-2018-0023-ver9-Gauna_1P.3d 08/06/18 9:16am Page 4

4 GAUNA ET AL.

FIG. 1. ANTHEPROT physicochemical profiles based on the primary structure of the core protein Peptide numbers are
the same in spot synthesis.

chemical control spots, numbered 23, 24, and 25, were added
as a repetition of peptides 9, 17, and 4, respectively. When the
membrane with peptides was exposed to a pool of four hepa-
titis C patients’ sera, antibody recognition was observed for
peptides 1, 3, 2, and 4, in decreasing order of signal intensity,
F2 c corresponding to the N-terminal end of the Core (Fig. 2).
Other sequences, such as spots 11, 13, 15, 16, 17, 18, and 24
(a repeat of spot 17) of nonstructural proteins, showed slight
antibody recognition. However, since we are looking for po-
tent antigens, these peptides were discarded for the time being.
No recognition was observed by selected sequences from the
structural protein E2, which is not surprising since its values of
hydrophilicity, solvent accessibility, and antigenicity were the
lowest ones of those observed in the immunoinformatic
analysis (data not shown). These results suggest that the Core
molecule, which is a highly conserved basic protein, is the
most antigenic under our study conditions.
This technique allowed us to reduce the selected peptides
to those corresponding to spots 1, 2, 3, and 4 for a larger
scale synthesis on resin, with and without incorporation of
stearic acid at the N-terminus, to be tested against a total of
22 hepatitis C patients’ sera by MABA.
Multiple antigen blot assay
Eight peptides were tested by MABA, corresponding to
sequences selected by Pepscan (spots 1–4) and their analogs
with stearic acid (Table 2). Peptide IMT-2052 (spot 1 in b T2

Table 2. Synthetic Peptides of Core Protein

With and Without Stearic Acid and Their
Frequency Recognition by Multiple
Antigen Blot Assay
Peptide Recognition
Spot Sequence (IMT-) by MABA
1 MSTNPKPQRKTKRNTNRRPQ 2052 22/22
Ste-MSTNPKPQRKTKRN 2065 16/22
TNRRPQ
2 RLGVRTTRKTSERSQPRGRR 2053 8/22
Ste-RLGVRTTRKTSERS 2066 8/22
QPRGRR
3 SQPRGRRQPIPKARRPEGRT 2054 13/22
Ste-SQPRGRRQPIPKARRPEGRT 2067 10/22
4 PSWGPTDPRRRSRNLGKVID 2055 0/22
Ste-PSWGPTDPRRRSRNLGKVID 2068 2/22
FIG. 2. Peptide reactivity against a pool of sera using IMT, Instituto de Medicina Tropical; MABA, multiple antigen
Pepscan methodology. blot assay.
VIM-2018-0023-ver9-Gauna_1P.3d 08/06/18 9:17am Page 5

USE OF SYNTHETIC PEPTIDES AND MABA IN HCV IMMUNODIAGNOSIS 5

Pepscan) showed antibody recognition by all 22 infected
patients’ sera included in this preliminary study. Most of
signals corresponding to this peptide were strong using both
enzymes, the anti-human IgG peroxidase and anti-human
IgG alkaline phosphatase. Its stearic acid analog to IMT-
F3 c 2052 resulted with recognition values of 16/22 (Fig. 3A, B).
Other peptides showed different degrees of antibody
recognition ranging from 13/22 to 0/22, as summarized
in Table 2.
Discussion
In previous studies, we have combined immunoinformatic
analysis, Pepscan methodology, and MABA technique in the
discovery of antigenic peptides for immunodiagnosis of hep-
atitis A infection (10). In this study, sequences of two structural
and two nonstructural proteins of HCV were analyzed in
combination with structural analysis of NS3 protein. Twenty-
two sequences, most of them of 20-mer, were selected for their
synthesis on a cellulose membrane.
During this Pepscan, we used amino acid solutions four
times less concentrated than previously used to synthesize
20-mer peptides on paper (10). Even when the lowest con-
centration of amino acid solutions was used, strong signals
were observed for spots 1–4, all of them from Core protein,
highlighting its relevance in the immunodiagnosis of hepa-
titis C infection.
It was precisely spot 1 that exhibited the strongest anti-
body recognition by Pepscan, and it was the peptide that
showed the highest sensitivity when synthesized and tested
by MABA. Peptide IMT-2052 had a recognition frequency
of 22/22, 100% of sensitivity, while sequences of peptides
IMT-2054, IMT-2053, and IMT-2055 showed sensitivities
of 59%, 30%, and 0% by MABA, respectively, which aligns
with the signal intensity observed by Pepscan. The 100%
IMT-2052 recognition frequency will lead us to confirm
these high values of sensitivity and specificity and to de-
termine the positive and negative predictive values through
epidemiological studies in the general population.
It has been reported that peptides containing bulky and
hydrophobic side chains may bind better to solid plastic
surfaces of certain enzyme-linked immunosorbent assay
(ELISA) plates (25). As mainly hydrophobic interactions are
responsible for adsorption to the surface of PVC plates as
well as NC paper, we may assume that the use of fatty acids
will provide the conjugate with the necessary hydropho-
bicity (11). In an attempt to optimize the MABA technique
and improve peptide adhesion to NC paper, we synthesized

FIG. 3. Antigenicity of synthetic peptides evaluated by MABA by sera of 22 hepatitis C patients and 5 negative controls
using (A) anti-human IgG peroxidase with Luminol SuperSignal West Pico Chemiluminescent Substrate and (B) anti-
human IgG alkaline phosphatase conjugates with NBT and BCIP as chromogenic substrates. No-SC and Control HS as
complementary controls were included. BCIP, bromo-4-chloro-3¢-indoly phosphate p-toluidine salt; HS, human sera;
MABA, multiple antigen blot assay; NBT, nitro blue tetrazolium chloride; No-SC, No-serum control.
VIM-2018-0023-ver9-Gauna_1P.3d 08/06/18 9:17am Page 6
6 GAUNA ET AL.
antigenic peptides with the incorporation of stearic acid at
N-terminus. However, we obtained better antibody recog-
nition with those peptides without the hydrophobic tail,
possibly due to a decrease in peptide solubility caused by
the incorporation of a fatty acid composed of 18 carbon
atoms, which is 2 carbons greater than palmitic acid that is
typically used. Fortunately, peptide IMT-2052 seems to
have good adhesion to NC paper and the incorporation of a
hydrophobic tail is not necessary to reach 100% sensitivity.
Other controls have been recently incorporated to the
MABA technique, such as the normal control HS. This
control consists in the immobilization of a serum as a source
of immunoglobulin in the first and last lanes of each strip,
which should always react in the presence of the secondary
antibody. In addition, a No-SC was used, which consists of a
strip not exposed to serum such that only a reaction with the
HS should be observed. These controls behaved as expected,
and all signals were reproducible with both enzymes.
MABA has been a useful technique, which permits us to
simultaneously test all peptides against the same serum at
the same time, reducing possible variations in study condi-
tions for different antigens. This low-cost technology re-
quires very small concentrations of antigens and very small
volumes of serum (6 mL), resulting in the identification of
molecules (peptides) at a low cost per patient ($ 0.006 per
MABA vs. $ 0.129 per conventional ELISA) (21). Further-
more, as the peptide synthesis becomes more widespread, its
costs decrease in comparison with the use and production of
purified or recombinant antigens.
Peptide IMT-2052 from the N-terminus of Core protein is
an interesting region of study for the design of inexpensive
diagnosis tests, which makes it more accessible for devel-
oping countries. It must be said that the major goal of our
research group is the development of a multidiagnosis kit
based on MABA technique and synthetic peptides, where
this sequence can join others of public health relevance to be
adapted and implemented for particular populations where
coinfections are frequent (5,20).
Conclusions
HCV infection is usually asymptomatic and it often re-
mains undiagnosed until serious liver damage has developed.
To prevent its transmission and the development of cirrhosis
and hepatic cancer, it is critical to have rapid, effective, and
inexpensive diagnostic methods that can be incorporated into
routine procedures in health centers in endemic areas. In this
work, we report the integration of a sequence analysis of the
HCV polyprotein subtype 1a (including structural and non-
structural viral proteins), followed by a spot synthesis
(Pepscan) of chosen peptides. This strategy allowed the se-
lection of four peptide sequences from the very conserved
Core protein, highlighting its relevance in the immunodiag-
nosis of viral hepatitis C infection. These four peptides
showed different levels of antigenic recognition by MABA,
with the peptide 1MSTNPKPQRKTKRNTNRRPQ20 (IMT-
2052) being the most antigenic, with a sensitivity and speci-
ficity of 100% utilizing sera from 22 hepatitis C patients.
Further epidemiological studies with a larger sample size are
necessary. Finally, we conclude that this peptide, with just
twenty residues, is a low-cost alternative for prevalence
studies and early diagnosis of hepatitis C infection.
Acknowledgments
This work has been partially supported by grants G-
2005000387, 201100413, 2007001425, F-2005000122, and
20122001311 from FONACIT-Venezuela, as well as Grand
Challenges Explorations grant funding number OPP1151004.
Author Disclosure Statement
No competing financial interests exist.
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Address correspondence to: b AU5
Dr. Oscar Noya
Sección de Biohelmintiasis
Instituto de Medicina Tropical
Facultad de Medicina
Universidad Central de Venezuela
Caracas
Venezuela
E-mail: noyaoo@yahoo.com
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