Stability of dried blood spots for HIV-1 drug resistance analysis

Anna C. Hearps, Claire E. Ryan, Lisa M Morris, Megan Plate, Vicki Greengrass, Suzanne M. Crowe

Research output: Contribution to journalArticleResearchpeer-review

20 Citations (Scopus)

Abstract

The wide scale application of dried blood spots (DBS) as a collection tool for low-cost HIV drug resistance testing requires a greater understanding of the accuracy of DBS for genotype analysis and the stability of DBS under various environmental conditions. Analysis of a 50μl DBS via a single amplicon, nested PCR-based in-house assay (the Burnet genotyping assay) showed an average nucleotide concordance of 98.9% with plasma samples, although only 65% of nucleotide mixtures detected in plasma were also detected within DBS. The analysis of three DBS resulted in the detection of a greater number of nucleotide mixtures (72 and 109 mixtures detected within one and three DBS, respectively, n = 10). Two DBS extraction protocols (silica particle; NucliSENS, bioMerieux and spin column extraction; High Pure, Roche) were assessed and found to be equivalent (79% and 84% recovery success respectively, n = 19). FTA Elute paper (Whatman) was an inferior DBS collection medium compared to Whatman 903 paper. DBS appeared relatively tolerant to multiple freeze/thaw cycles, with 79% of DBS subjected to ten freeze/thaw cycles successfully amplified compared to 93% of DBS defrosted once (n = 14). High temperature (37°C) and high humidity (>90%) substantially impaired DBS recovery within two weeks of storage (38%, n = 8), whilst storage at -20°C or 4°C adequately preserved DBS for this period (100% recovery, n = 8). Therefore, whilst DBS are suitable for HIV drug resistance surveillance, the use of multiple DBS may be required to ensure accurate detection of minor HIV quasispecies and shortterm storage of samples at either 4°C or -20°C is recommended.

Original languageEnglish
Pages (from-to)134-140
Number of pages7
JournalCurrent HIV Research
Volume8
Issue number2
DOIs
Publication statusPublished - Mar 2010
Externally publishedYes

Keywords

  • Antiretroviral therapy
  • Dried blood spot
  • HIV drug resistance
  • HIV genotyping

Cite this

Hearps, Anna C. ; Ryan, Claire E. ; Morris, Lisa M ; Plate, Megan ; Greengrass, Vicki ; Crowe, Suzanne M. / Stability of dried blood spots for HIV-1 drug resistance analysis. In: Current HIV Research. 2010 ; Vol. 8, No. 2. pp. 134-140.
@article{f5b5beaf801f41c98eab7ce33b01abfc,
title = "Stability of dried blood spots for HIV-1 drug resistance analysis",
abstract = "The wide scale application of dried blood spots (DBS) as a collection tool for low-cost HIV drug resistance testing requires a greater understanding of the accuracy of DBS for genotype analysis and the stability of DBS under various environmental conditions. Analysis of a 50μl DBS via a single amplicon, nested PCR-based in-house assay (the Burnet genotyping assay) showed an average nucleotide concordance of 98.9{\%} with plasma samples, although only 65{\%} of nucleotide mixtures detected in plasma were also detected within DBS. The analysis of three DBS resulted in the detection of a greater number of nucleotide mixtures (72 and 109 mixtures detected within one and three DBS, respectively, n = 10). Two DBS extraction protocols (silica particle; NucliSENS, bioMerieux and spin column extraction; High Pure, Roche) were assessed and found to be equivalent (79{\%} and 84{\%} recovery success respectively, n = 19). FTA Elute paper (Whatman) was an inferior DBS collection medium compared to Whatman 903 paper. DBS appeared relatively tolerant to multiple freeze/thaw cycles, with 79{\%} of DBS subjected to ten freeze/thaw cycles successfully amplified compared to 93{\%} of DBS defrosted once (n = 14). High temperature (37°C) and high humidity (>90{\%}) substantially impaired DBS recovery within two weeks of storage (38{\%}, n = 8), whilst storage at -20°C or 4°C adequately preserved DBS for this period (100{\%} recovery, n = 8). Therefore, whilst DBS are suitable for HIV drug resistance surveillance, the use of multiple DBS may be required to ensure accurate detection of minor HIV quasispecies and shortterm storage of samples at either 4°C or -20°C is recommended.",
keywords = "Antiretroviral therapy, Dried blood spot, HIV drug resistance, HIV genotyping",
author = "Hearps, {Anna C.} and Ryan, {Claire E.} and Morris, {Lisa M} and Megan Plate and Vicki Greengrass and Crowe, {Suzanne M.}",
year = "2010",
month = "3",
doi = "10.2174/157016210790442740",
language = "English",
volume = "8",
pages = "134--140",
journal = "Current HIV Research",
issn = "1570-162X",
publisher = "Bentham Science Publishers",
number = "2",

}

Stability of dried blood spots for HIV-1 drug resistance analysis. / Hearps, Anna C.; Ryan, Claire E.; Morris, Lisa M; Plate, Megan; Greengrass, Vicki; Crowe, Suzanne M.

In: Current HIV Research, Vol. 8, No. 2, 03.2010, p. 134-140.

Research output: Contribution to journalArticleResearchpeer-review

TY - JOUR

T1 - Stability of dried blood spots for HIV-1 drug resistance analysis

AU - Hearps, Anna C.

AU - Ryan, Claire E.

AU - Morris, Lisa M

AU - Plate, Megan

AU - Greengrass, Vicki

AU - Crowe, Suzanne M.

PY - 2010/3

Y1 - 2010/3

N2 - The wide scale application of dried blood spots (DBS) as a collection tool for low-cost HIV drug resistance testing requires a greater understanding of the accuracy of DBS for genotype analysis and the stability of DBS under various environmental conditions. Analysis of a 50μl DBS via a single amplicon, nested PCR-based in-house assay (the Burnet genotyping assay) showed an average nucleotide concordance of 98.9% with plasma samples, although only 65% of nucleotide mixtures detected in plasma were also detected within DBS. The analysis of three DBS resulted in the detection of a greater number of nucleotide mixtures (72 and 109 mixtures detected within one and three DBS, respectively, n = 10). Two DBS extraction protocols (silica particle; NucliSENS, bioMerieux and spin column extraction; High Pure, Roche) were assessed and found to be equivalent (79% and 84% recovery success respectively, n = 19). FTA Elute paper (Whatman) was an inferior DBS collection medium compared to Whatman 903 paper. DBS appeared relatively tolerant to multiple freeze/thaw cycles, with 79% of DBS subjected to ten freeze/thaw cycles successfully amplified compared to 93% of DBS defrosted once (n = 14). High temperature (37°C) and high humidity (>90%) substantially impaired DBS recovery within two weeks of storage (38%, n = 8), whilst storage at -20°C or 4°C adequately preserved DBS for this period (100% recovery, n = 8). Therefore, whilst DBS are suitable for HIV drug resistance surveillance, the use of multiple DBS may be required to ensure accurate detection of minor HIV quasispecies and shortterm storage of samples at either 4°C or -20°C is recommended.

AB - The wide scale application of dried blood spots (DBS) as a collection tool for low-cost HIV drug resistance testing requires a greater understanding of the accuracy of DBS for genotype analysis and the stability of DBS under various environmental conditions. Analysis of a 50μl DBS via a single amplicon, nested PCR-based in-house assay (the Burnet genotyping assay) showed an average nucleotide concordance of 98.9% with plasma samples, although only 65% of nucleotide mixtures detected in plasma were also detected within DBS. The analysis of three DBS resulted in the detection of a greater number of nucleotide mixtures (72 and 109 mixtures detected within one and three DBS, respectively, n = 10). Two DBS extraction protocols (silica particle; NucliSENS, bioMerieux and spin column extraction; High Pure, Roche) were assessed and found to be equivalent (79% and 84% recovery success respectively, n = 19). FTA Elute paper (Whatman) was an inferior DBS collection medium compared to Whatman 903 paper. DBS appeared relatively tolerant to multiple freeze/thaw cycles, with 79% of DBS subjected to ten freeze/thaw cycles successfully amplified compared to 93% of DBS defrosted once (n = 14). High temperature (37°C) and high humidity (>90%) substantially impaired DBS recovery within two weeks of storage (38%, n = 8), whilst storage at -20°C or 4°C adequately preserved DBS for this period (100% recovery, n = 8). Therefore, whilst DBS are suitable for HIV drug resistance surveillance, the use of multiple DBS may be required to ensure accurate detection of minor HIV quasispecies and shortterm storage of samples at either 4°C or -20°C is recommended.

KW - Antiretroviral therapy

KW - Dried blood spot

KW - HIV drug resistance

KW - HIV genotyping

UR - http://www.scopus.com/inward/record.url?scp=77949716973&partnerID=8YFLogxK

U2 - 10.2174/157016210790442740

DO - 10.2174/157016210790442740

M3 - Article

C2 - 20163343

AN - SCOPUS:77949716973

VL - 8

SP - 134

EP - 140

JO - Current HIV Research

JF - Current HIV Research

SN - 1570-162X

IS - 2

ER -