TY - JOUR
T1 - Next-generation viral RNA/DNA in situ hybridization applications in human immunodeficiency virus/simian immunodeficiency virus research
AU - Brands, Catherine
AU - Morcock, David
AU - Estes, Jacob
AU - Deleage, Claire
N1 - Funding Information:
This project has been funded in whole with Federal funds from the National Cancer Institute, National Institutes of Health, under Contract No. HHSN261200800001E and by the Oregon National Primate Research Center NIH grant award P51OD011092 (J.D.E). The content of this publication does not necessarily reflect the views or policies of the Department of Health and Human Services, nor does mention of trade names, commercial products, or organizations imply endorsement by the U.S. Government. The duplex was developed with the help of Advanced Cell Diagnostics.
Publisher Copyright:
© 2020 JoVE Creative Commons Attribution-NonCommercial-NoDerivs 3.0 Unported License.
PY - 2020/6
Y1 - 2020/6
N2 - In situ hybridization is a powerful technique to identify specific RNA or DNA sequences within individual cells in tissue sections, providing important insights into physiological processes and disease pathogenesis. In situ hybridization (ISH) has been used for many years to assess the location of cells infected by viruses, but recently a next-generation ISH approach was developed with a unique probe design strategy that allows simultaneous signal amplification and background suppression to achieve single-molecule visualization while preserving tissue morphology. This next-generation ISH is based on an approach like branched PCR, but performed in situ and is more facile, sensitive, and reproducible than classical ISH methods or in situ PCR approaches in routinely detecting RNA or DNA in formalin-fixed paraffin embedded (FFPE) tissues. For the last several years our laboratory has been applying this ISH platform for the detection of human immunodeficiency (HIV) and simian immunodeficiency (SIV) viral RNA (vRNA) and/or viral DNA (vDNA) positive cells within a multitude of FFPE tissues. With this detailed technical manuscript, we would like to share our knowledge and advice with all individuals interested in using next-generation ISH in their research.
AB - In situ hybridization is a powerful technique to identify specific RNA or DNA sequences within individual cells in tissue sections, providing important insights into physiological processes and disease pathogenesis. In situ hybridization (ISH) has been used for many years to assess the location of cells infected by viruses, but recently a next-generation ISH approach was developed with a unique probe design strategy that allows simultaneous signal amplification and background suppression to achieve single-molecule visualization while preserving tissue morphology. This next-generation ISH is based on an approach like branched PCR, but performed in situ and is more facile, sensitive, and reproducible than classical ISH methods or in situ PCR approaches in routinely detecting RNA or DNA in formalin-fixed paraffin embedded (FFPE) tissues. For the last several years our laboratory has been applying this ISH platform for the detection of human immunodeficiency (HIV) and simian immunodeficiency (SIV) viral RNA (vRNA) and/or viral DNA (vDNA) positive cells within a multitude of FFPE tissues. With this detailed technical manuscript, we would like to share our knowledge and advice with all individuals interested in using next-generation ISH in their research.
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U2 - 10.3791/60318
DO - 10.3791/60318
M3 - Article
C2 - 32628155
AN - SCOPUS:85088210852
SN - 1940-087X
VL - 2020
SP - 1
EP - 13
JO - Journal of visualized experiments : JoVE
JF - Journal of visualized experiments : JoVE
IS - 160
M1 - e60318
ER -