In situ polymerase chain reaction amplification of HIV-1 DNA in brain tissue

P M Strappe; T H Wang; C A McKenzie; S Lowrie; P Simmonds; J E Bell

doi:10.1016/S0166-0934(97)00156-0

In situ polymerase chain reaction amplification of HIV-1 DNA in brain tissue

P M Strappe, T H Wang, C A McKenzie, S Lowrie, P Simmonds, J E Bell

Royal (Dick) School of Veterinary Studies

Research output: Contribution to journal › Article › peer-review

Abstract

A direct in situ polymerase chain reaction (IS-PCR) assay is described for the detection of HIV-1 proviral DNA in formalin fixed paraffin embedded brain tissue. Biotin-16-dUTP is incorporated during the PCR process and microwave pretreatment of tissue sections ensures that no non-specific incorporation into damaged or nicked genomic DNA occurs. Two methods are compared to detect the biotinylated amplified product, the use of an avidin-biotin-alkaline phosphatase complex (ABC) and the application of tyramide signal amplification (TSA) which allows both chromogenic and fluorescence detection. TSA detection enhances the sensitivity of IS-PCR, permitting fewer PCR cycles and preserving tissue morphology.

Original language	English
Pages (from-to)	119-27
Number of pages	9
Journal	Journal of Virological Methods
Volume	70
Issue number	2
DOIs	https://doi.org/10.1016/S0166-0934(97)00156-0
Publication status	Published - Feb 1998

Keywords / Materials (for Non-textual outputs)

Brain
DNA, Viral
Formaldehyde
HIV-1
Humans
Microwaves
Paraffin Embedding
Polymerase Chain Reaction
Sensitivity and Specificity
Tissue Fixation

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10.1016/S0166-0934(97)00156-0

Cite this

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title = "In situ polymerase chain reaction amplification of HIV-1 DNA in brain tissue",

abstract = "A direct in situ polymerase chain reaction (IS-PCR) assay is described for the detection of HIV-1 proviral DNA in formalin fixed paraffin embedded brain tissue. Biotin-16-dUTP is incorporated during the PCR process and microwave pretreatment of tissue sections ensures that no non-specific incorporation into damaged or nicked genomic DNA occurs. Two methods are compared to detect the biotinylated amplified product, the use of an avidin-biotin-alkaline phosphatase complex (ABC) and the application of tyramide signal amplification (TSA) which allows both chromogenic and fluorescence detection. TSA detection enhances the sensitivity of IS-PCR, permitting fewer PCR cycles and preserving tissue morphology.",

keywords = "Brain, DNA, Viral, Formaldehyde, HIV-1, Humans, Microwaves, Paraffin Embedding, Polymerase Chain Reaction, Sensitivity and Specificity, Tissue Fixation",

author = "Strappe, {P M} and Wang, {T H} and McKenzie, {C A} and S Lowrie and P Simmonds and Bell, {J E}",

year = "1998",

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doi = "10.1016/S0166-0934(97)00156-0",

language = "English",

volume = "70",

pages = "119--27",

journal = "Journal of Virological Methods",

issn = "0166-0934",

publisher = "Elsevier",

number = "2",

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TY - JOUR

T1 - In situ polymerase chain reaction amplification of HIV-1 DNA in brain tissue

AU - Strappe, P M

AU - Wang, T H

AU - McKenzie, C A

AU - Lowrie, S

AU - Simmonds, P

AU - Bell, J E

PY - 1998/2

Y1 - 1998/2

N2 - A direct in situ polymerase chain reaction (IS-PCR) assay is described for the detection of HIV-1 proviral DNA in formalin fixed paraffin embedded brain tissue. Biotin-16-dUTP is incorporated during the PCR process and microwave pretreatment of tissue sections ensures that no non-specific incorporation into damaged or nicked genomic DNA occurs. Two methods are compared to detect the biotinylated amplified product, the use of an avidin-biotin-alkaline phosphatase complex (ABC) and the application of tyramide signal amplification (TSA) which allows both chromogenic and fluorescence detection. TSA detection enhances the sensitivity of IS-PCR, permitting fewer PCR cycles and preserving tissue morphology.

AB - A direct in situ polymerase chain reaction (IS-PCR) assay is described for the detection of HIV-1 proviral DNA in formalin fixed paraffin embedded brain tissue. Biotin-16-dUTP is incorporated during the PCR process and microwave pretreatment of tissue sections ensures that no non-specific incorporation into damaged or nicked genomic DNA occurs. Two methods are compared to detect the biotinylated amplified product, the use of an avidin-biotin-alkaline phosphatase complex (ABC) and the application of tyramide signal amplification (TSA) which allows both chromogenic and fluorescence detection. TSA detection enhances the sensitivity of IS-PCR, permitting fewer PCR cycles and preserving tissue morphology.

KW - Brain

KW - DNA, Viral

KW - Formaldehyde

KW - HIV-1

KW - Humans

KW - Microwaves

KW - Paraffin Embedding

KW - Polymerase Chain Reaction

KW - Sensitivity and Specificity

KW - Tissue Fixation

U2 - 10.1016/S0166-0934(97)00156-0

DO - 10.1016/S0166-0934(97)00156-0

M3 - Article

C2 - 9562406

SN - 0166-0934

VL - 70

SP - 119

EP - 127

JO - Journal of Virological Methods

JF - Journal of Virological Methods

IS - 2

ER -

In situ polymerase chain reaction amplification of HIV-1 DNA in brain tissue

Abstract

Keywords / Materials (for Non-textual outputs)

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