Aug 28, 2020

Public workspaceCapture and concentration of SARS-CoV-2 and other respiratory viruses from VTM / UTM samples using Magnetic Nanotrap® particles for direct RNA extraction

This protocol is a draft, published without a DOI.
  • Ben Lepene1,
  • Anurag Patnaik1,
  • Robert Barclay1
  • 1Ceres Nanosciences, Inc
  • Coronavirus Method Development Community
  • Nanotrap Applications
Ben Lepene
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Protocol Citation: Ben Lepene, Anurag Patnaik, Robert Barclay 2020. Capture and concentration of SARS-CoV-2 and other respiratory viruses from VTM / UTM samples using Magnetic Nanotrap® particles for direct RNA extraction. protocols.io https://protocols.io/view/capture-and-concentration-of-sars-cov-2-and-other-bh2rj8d6
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License,  which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: June 29, 2020
Last Modified: August 28, 2020
Protocol Integer ID: 38705
Keywords: virus , capture, extraction, concentration, VTM, UTM, Nanotrap, viral, Nanoscience, Ceres, SARS-CoV-2, COVID-19,
Abstract
This protocol provides a method for Magnetic Nanotrap® particle-based capture and concentration of viruses from Viral transport media and Universal transport media samples.
Materials
MATERIALS
ReagentTriton(R) X-100 100mlPromegaCatalog #H5142
ReagentDynaMag™-2 Magnet Life TechnologiesCatalog #12321D
ReagentMicroAmp™ Optical 96-Well Reaction Plate with Barcode & Optical Adhesive FilmsThermo FisherCatalog #4314320
ReagentRT-PCR Grade WaterThermo FisherCatalog #AM9935
ReagentMini Vortex MixerCatalog #M10101001
ReagentNanotrap Magnetic Virus Particles (10)Ceres NanoCatalog #44202
ReagentViral Transport Media (VTM)
ReagentUniversal Transport Media (UTM)
ReagentMicrocentrifuge Tubes
ReagentRT-PCR Kit
Reagent1x PBS without Calcium & MagnesiumLonzaCatalog #17-516Q
Materials and equipment:
Magnetic Nanotrap® particles
Magnetic separator
Extraction buffer - 0.5% Triton X-100 in PCR grade water
Vortex
Roche Lightcycler 96*
Optical Adhesive Film
96 Well PCR plate
*Alternative PCR kits, real-time PCR machines, and positive controls can be utilized with this protocol*
Safety warnings
Please refer to Safety Data Sheets (SDS) for health and environmental hazards.
Virus Capture and RNA Extraction
Virus Capture and RNA Extraction
20m
20m
Add Amount1 mL Viral transport media (VTM) or Universal transport media (UTM) sample to a microcentrifuge tube.
Note
500 µL of the sample can be used if 1 mL is not available.


30s
Add Amount100 µL of Magnetic Nanotrap® particles to the sample.

30s
Pipetting
Incubate samples with Magnetic Nanotrap® particles at TemperatureRoom temperature for Duration00:10:00 .

10m
Incubation
Use a magnetic rack to separate the Magnetic Nanotrap® particles from the sample.Duration00:01:00

1m
Discard the supernatant carefully without disturbing the pellet.
30s
Add Amount500 µL of 1X PBS to the pellet and resuspend to wash.

1m
Wash
Use a magnetic rack to separate the Magnetic Nanotrap® particles from the sample.Duration00:01:00

1m
Discard the supernatant carefully without disturbing the pellet. If required - use a smaller pipette to remove any residual PBS.
30s
Critical
Resuspend particle pellet in Amount50 µL of extraction buffer .

30s
Pipetting
Heat samples at Temperature95 °C for Duration00:05:00 .
Note
This step can be performed on a heat block or thermocycler.


5m
Critical
Use a magnetic rack to separate the Magnetic Nanotrap® particles from the sample.Duration00:00:30

45s
Collect the supernatant. The sample is ready for analysis.
30s
RT-PCR Reaction
RT-PCR Reaction
Use any SARS-CoV-2 RT-PCR detection kit. Follow manufacturer instructions to set up the RT- PCR
PCR