ChloroquineV1, France, Analysis, bibRecord, 000143

Drying anti-malarial drugs in vitro tests to outsource SYBR green assays

Identifieur interne : 000143 ( France/Analysis ); précédent : 000142; suivant : 000144

Drying anti-malarial drugs in vitro tests to outsource SYBR green assays

Auteurs : Karim-Frédéric Traore [France] ; Adeline Lavoignat [France] ; Guillaume Bonnot [France] ; Fatimata Sow [France] ; Giuliana C. Bess [France] ; Marjorie Chavant [France] ; Frederick Gay [France] ; Ogobara Doumbo [Mali] ; Stephane Picot [France]

Source :

Malaria Journal [ 1475-2875 ] ; 2015-02-21.

RBID : Hal:hal-01121828

English descriptors

mix :
- Drug resistance, Ex vivo, In vitro, Malaria, SYBR green.

Abstract

BackgroundMeasurement of anti-malarial drug efficacy and resistance relies mainly on in vivo clinical trials, in vitro/ex vivo assays and molecular markers detection. The existing in vitro/ex vivo assays, in particular those that are using non-radioactive devices, need to be standardized and adapted to field conditions. SYBR Green assay offers a rapid and cheap alternative to other in vitro assays, but it requires tools not commonly available in field laboratories. Here is described a modified SYBR green I protocol to perform the parasite growth test with blood samples in endemic areas, followed later by the SYBR green fluorescence assay performed at a specialized laboratory level.MethodsIn vitro susceptibility of Plasmodium falciparum clones HB3, 3D7, W2 and 7G8 to chloroquine (CQ), dihydroartemisinin (DHA), pyronaridine (PYD) and piperaquine (PPQ) was tested. Fresh isolates of P. falciparum from imported malaria cases were collected for ex vivo assays. The parasite suspension was added in 96-well plates predosed with anti-malarial drugs and incubated for 72 hours at 37°C, 5% CO2. SYBR green I protocol was modified to dry the plates after freeze-thawed process to mimic storage and shipping conditions. The plates were rehydrated with 200 μl of complete RPMI medium for fluorescence assay.ResultsThere were no significant differences in IC50 values of CQ, DHA, PYD and PPQ, determined by the modified protocol, compared to standard protocol. Longer storage did not affect the IC50 values.ConclusionThe SYBR green I modified protocol produced reliable results and could be a suitable method for in vitro/ex vivo assays in field.

Url:

https://hal.sorbonne-universite.fr/hal-01121828

DOI: 10.1186/s12936-015-0600-z

Affiliations:

France, Mali

Links toward previous steps (curation, corpus...)

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to stream Main, to step Merge: 000E54
to stream Main, to step Curation: 000E53
to stream Main, to step Exploration: 000E53
to stream France, to step Extraction: 000143

Links to Exploration step

Hal:hal-01121828

Le document en format XML

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<tutelle active="#struct-454982" type="direct"><org type="institution" xml:id="struct-454982" status="VALID">          <orgName>CHU Saint-Antoine [APHP]</orgName>
          <desc>            <address>              <addrLine>184, rue du Faubourg Saint-Antoine 75571 Paris cedex 12</addrLine>
              <country key="FR"></country>
            </address>
            <ref type="url">http://www.aphp.fr/contenu/hopital-saint-antoine-1</ref>
          </desc>
        </org>
</tutelle>
</tutelles>
</hal:affiliation>
<country>France</country>
</affiliation>
</author>
<author><name sortKey="Doumbo, Ogobara" sort="Doumbo, Ogobara" uniqKey="Doumbo O" first="Ogobara" last="Doumbo">Ogobara Doumbo</name>
<affiliation wicri:level="1"><hal:affiliation type="laboratory" xml:id="struct-197154" status="VALID">          <orgName>Malaria Research and Training Center [Bamako, Mali]</orgName>
          <desc>            <address>              <country key="ML"></country>
            </address>
          </desc>
          <listRelation>            <relation active="#struct-218026" type="direct"></relation>
          </listRelation>
        <tutelles><tutelle active="#struct-218026" type="direct"><org type="institution" xml:id="struct-218026" status="VALID">          <orgName>Université de Bamako</orgName>
          <desc>            <address>              <addrLine>BP E.2528 Bamako</addrLine>
              <country key="ML"></country>
            </address>
          </desc>
        </org>
</tutelle>
</tutelles>
</hal:affiliation>
<country>Mali</country>
</affiliation>
</author>
<author><name sortKey="Picot, Stephane" sort="Picot, Stephane" uniqKey="Picot S" first="Stephane" last="Picot">Stephane Picot</name>
<affiliation wicri:level="1"><hal:affiliation type="laboratory" xml:id="struct-230740" status="INCOMING">          <orgName>Malaria Research Unit</orgName>
          <desc>            <address>              <addrLine>Lyon</addrLine>
              <country key="FR"></country>
            </address>
          </desc>
          <listRelation>            <relation name="UMR5246" active="#struct-441569" type="direct"></relation>
          </listRelation>
        <tutelles><tutelle name="UMR5246" active="#struct-441569" type="direct"><org type="institution" xml:id="struct-441569" status="VALID">          <idno type="IdRef">02636817X</idno>
          <idno type="ISNI">0000000122597504</idno>
          <orgName>Centre National de la Recherche Scientifique</orgName>
          <orgName type="acronym">CNRS</orgName>
          <date type="start">1939-10-19</date>
          <desc>            <address>              <country key="FR"></country>
            </address>
            <ref type="url">http://www.cnrs.fr/</ref>
          </desc>
        </org>
</tutelle>
</tutelles>
</hal:affiliation>
<country>France</country>
</affiliation>
</author>
</analytic>
<idno type="DOI">10.1186/s12936-015-0600-z</idno>
<series><title level="j">Malaria Journal</title>
<idno type="ISSN">1475-2875</idno>
<imprint><date type="datePub">2015-02-21</date>
</imprint>
</series>
</biblStruct>
</sourceDesc>
</fileDesc>
<profileDesc><textClass><keywords scheme="mix" xml:lang="en"><term>Drug resistance</term>
<term>Ex vivo</term>
<term>In vitro</term>
<term>Malaria</term>
<term>SYBR green</term>
</keywords>
</textClass>
</profileDesc>
</teiHeader>
<front><div type="abstract" xml:lang="en">              <p>BackgroundMeasurement of anti-malarial drug efficacy and resistance relies mainly on in vivo clinical trials, in vitro/ex vivo assays and molecular markers detection. The existing in vitro/ex vivo assays, in particular those that are using non-radioactive devices, need to be standardized and adapted to field conditions. SYBR Green assay offers a rapid and cheap alternative to other in vitro assays, but it requires tools not commonly available in field laboratories. Here is described a modified SYBR green I protocol to perform the parasite growth test with blood samples in endemic areas, followed later by the SYBR green fluorescence assay performed at a specialized laboratory level.MethodsIn vitro susceptibility of Plasmodium falciparum clones HB3, 3D7, W2 and 7G8 to chloroquine (CQ), dihydroartemisinin (DHA), pyronaridine (PYD) and piperaquine (PPQ) was tested. Fresh isolates of P. falciparum from imported malaria cases were collected for ex vivo assays. The parasite suspension was added in 96-well plates predosed with anti-malarial drugs and incubated for 72 hours at 37°C, 5% CO2. SYBR green I protocol was modified to dry the plates after freeze-thawed process to mimic storage and shipping conditions. The plates were rehydrated with 200 μl of complete RPMI medium for fluorescence assay.ResultsThere were no significant differences in IC50 values of CQ, DHA, PYD and PPQ, determined by the modified protocol, compared to standard protocol. Longer storage did not affect the IC50 values.ConclusionThe SYBR green I modified protocol produced reliable results and could be a suitable method for in vitro/ex vivo assays in field.</p>
            </div>
</front>
</TEI>
<affiliations><list><country><li>France</li>
<li>Mali</li>
</country>
</list>
<tree><country name="France"><noRegion><name sortKey="Traore, Karim Frederic" sort="Traore, Karim Frederic" uniqKey="Traore K" first="Karim-Frédéric" last="Traore">Karim-Frédéric Traore</name>
</noRegion>
<name sortKey="Bess, Giuliana C" sort="Bess, Giuliana C" uniqKey="Bess G" first="Giuliana C" last="Bess">Giuliana C. Bess</name>
<name sortKey="Bonnot, Guillaume" sort="Bonnot, Guillaume" uniqKey="Bonnot G" first="Guillaume" last="Bonnot">Guillaume Bonnot</name>
<name sortKey="Chavant, Marjorie" sort="Chavant, Marjorie" uniqKey="Chavant M" first="Marjorie" last="Chavant">Marjorie Chavant</name>
<name sortKey="Gay, Frederick" sort="Gay, Frederick" uniqKey="Gay F" first="Frederick" last="Gay">Frederick Gay</name>
<name sortKey="Lavoignat, Adeline" sort="Lavoignat, Adeline" uniqKey="Lavoignat A" first="Adeline" last="Lavoignat">Adeline Lavoignat</name>
<name sortKey="Picot, Stephane" sort="Picot, Stephane" uniqKey="Picot S" first="Stephane" last="Picot">Stephane Picot</name>
<name sortKey="Sow, Fatimata" sort="Sow, Fatimata" uniqKey="Sow F" first="Fatimata" last="Sow">Fatimata Sow</name>
</country>
<country name="Mali"><noRegion><name sortKey="Doumbo, Ogobara" sort="Doumbo, Ogobara" uniqKey="Doumbo O" first="Ogobara" last="Doumbo">Ogobara Doumbo</name>
</noRegion>
</country>
</tree>
</affiliations>
</record>

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Drying anti-malarial drugs in vitro tests to outsource SYBR green assays

Drying anti-malarial drugs in vitro tests to outsource SYBR green assays

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