Target Capture; Transcription-Mediated Amplification (Tma) - Procleix TIGRIS Operator's Manual

Target Capture

The Target Capture process isolates the viral RNA and DNA from plasma specimens and
purifies it for amplification.
1. The Sample Pipettor combines each sample with working Target Capture Reagent
(wTCR) in one tube of a Multi-Tube Unit (MTU). The right mixer then mixes the sample
and wTCR. In the PROCLEIX ULTRIO Plus
Mixer, where Target Enhancer Reagent is added to each reaction tube to enhance the
disruption of the HBV viral particles. Finally, the MTU is moved to the Anneal Incubator for
the Target Capture reaction.
a) TCR solubilizes the viral envelope, denatures proteins, and releases viral genomic
RNA/DNA.
b) Oligonucleotides called "capture oligonucleotides" hybridize to the target RNA/DNA
sequences from the lysed virus.
c) The capture oligonucleotide and target RNA/DNA complexes attach to magnetic
particles by their tail sequence.
2. The MTU enters a Magnetic Wash Station (MWS):
a) In the MWS, magnets pull the magnetic particles, bound to target RNA/DNA
complexes, to the sides of the tubes.
b) Aspirators in the MWS aspirate denatured plasma components. Removal of this
debris purifies the target RNA/DNA for amplification.
c) The MWS adds Wash Solution to each tube of the MTU, mixes the contents,
reapplies the magnet, and aspirates the contents. This wash step and magnetic
binding step is repeated for each reaction tube.
d) Liquid Level Sensing (LLS) verifies Wash Solution dispense and aspiration via
conductive tiplets.
3. When the magnetic wash cycle is complete, the target RNA/DNA is pure enough to be
amplified by Transcription-Mediated Amplification.

Transcription-Mediated Amplification (TMA)

Transcription-Mediated Amplification (TMA) uses two enzymes (Reverse Transcriptase and RNA
Polymerase) to produce billions of copies of RNA amplified product from the purified target.
The TMA process takes approximately one hour.
1. Oil is dispensed into the MTU, then the Amplification Pipettor dispenses Amplification
Reagent. The left mixer mixes the reagents.
a) Oil helps to prevent evaporation and the generation of aerosols, which can
contaminate other samples.
b) Amplification Reagent contains dNTPs, NTPs and oligonucleotide primers. These
primers help the two enzymes create copies of the target RNA/DNA.
2. The MTU moves to the Anneal Incubator. Incubation promotes the binding of the primers
to the target RNA/DNA.
3. The MTU moves to the Transition Incubator, then to the Amplification Incubator, where the
Amplification Pipettor dispenses Enzyme Reagent into the MTU.
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PROCLEIX TIGRIS System Operator's Manual
Volume I: Operating the Instrument
I
NTRODUCTION AND
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Assay, the MTU is then moved to the Left
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RODUCT ESCRIPTION
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