Innovative Uses of RNAscope

Read about reseachers who take innovative approaches with RNAscope® protocols to advance their research. If you would like to try one of these customer-demonstrated applications, please note that they are non-standard applications and therefore, a careful read through of the publications/materials is highly recommended. Our technical support team is here and will make best efforts to support you.

Customer demonstrated application:
SIV/HIV RNA & DNA detection

ACD congratulates the team of scientists, led by Jacob D. Estes on their publication Defining HIV and SIV Reservoirs in Lymphoid Tissues DOI 10.20411/pai.v1i1.100.

The authors aimed to track and discriminate HIV-1/SIV viral reservoirs within tissue compartments and thus applied a 'specific and sensitive next-generation in situ hybridization approach.' The authors demonstrated:

1) that an optimized next-generation ISH platform RNAscope Assay for the rapid detection of vRNA (with results obtained within 1 day) has sufficient sensitivity to reliably detect single virions in B cell follicles (BCF) in FFPE tissue sections,

2) that an approach for the detection of vDNA in situ (referred to as DNAscope) reliably and readily detects vDNA+ cells, and

3) that they have developed an in situ method to simultaneously visualize vRNA and vDNA in the same tissue section and thereby identify transcriptionally latent infections (vDNA+/vRNA- cells) in lymphoid tissues.

The team of researchers then applied “these new, highly sensitive in situ hybridization approaches" to lymphoid tissues samples from macaques prior to and during combination anti-retroviral therapy (cART) and documented the importance of BCFs in active, latent and persistent infections during treatment. Authors concluded that “these data underscore the utility of new and sensitive ISH tools that can provide additional insight into viral persistence, reservoir establishment, tissue compartmentalization and reservoir phenotype in the local in vivo tissue environment.”

Read the Publication

Contact ACD to get started

Visualization of viral DNA as described above is a non-standard, modified RNAscope® protocol.  Please familiarize yourself with the methodology described. The ACD technical support team will provide the best support for you.  Please note, as yet, there are no experimental data demonstrating the extension of this protocol to other species, other probes or other tissue types not described in the publication.  

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What to order from ACD

Your account executive will ensure completeness of your order.  Standard accessories needed for any of the three protocols below include:

1. ACD HybEZ Hydridization System with ACD EZBatch Cat. No. 321461 110V or Cat. No. 321462 220V

2. ImmEdge Hydrophobic Barrier Pen Cat. No. 310018

3. Control probes:  Negative control probe and species-specific positive control probes are also required 

 

To detect and visualize HIV-1 AND SIV viral RNA in situ, the authors used standard RNAscope Assay Protocol and anti-sense probes outlined in Supplemental Table 2. Additionally the following materials from ACD are required:

1.  RNAscope 2.5 HD Red Kit: Cat. No. 322350 

2. EcoMount Mounting Medium Cat. No. 320409

 

To detect and visualize HIV-1 AND SIV viral DNA in situ, the authors used modified RNAscope Assay Protocol and sense probes outlined in Supplemental Table 2. Additionally the following materials from ACD are required:

1.RNAscope 2.5 HD Brown Kit Cat. No. 322300

 

To detect and visualize both viral RNA and viral DNA in situ, in a single assay the authors used modified RNAscope Assay Protocol both sense and anti-sense probes outlined in Supplemental Table 3. The following materials from ACD are also required:

1. RNAscope 2.5 HD Duplex Kit Cat. No. 322430

2. VectaMount Permanent Mounting Medium Cat. No.321584

 

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