Protocol Citation: Daniel Benner, Ozlem Yaren 2020. FireLAMP Protocol. protocols.io https://dx.doi.org/10.17504/protocols.io.bkz7kx9n
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: September 07, 2020
Last Modified: September 08, 2020
Protocol Integer ID: 41759
Materials
| Reagent or Consumable | Supplier | Catalog # | Cost per item (e.g., cost of a tube of enzyme) | |
| Text | Text | Numeric | Numeric | |
| SARS-CoV-2 Primers | IDT | Custom | $100.00 | |
| SARS-CoV-2 Probes | IDT | Custom | $280.00 | |
| Internal Control Primers | IDT | Custom | $100.00 | |
| Internal Control Probes | IDT | Custom | $280.00 | |
| WarmStart® LAMP Kit (DNA & RNA) | NEB | E1700L | $844.00 | |
| Antarctic thermolabile UDG | NEB | M0372L | $310.00 | |
| RNase Inhibitor, Murine | NEB | M0314L | $286.00 | |
| dUTP | NEB | N0459S | $60.00 | |
| Elution solution | Firebird Biomolecular Sciences, LLC | Custom | $0.50 | |
| 1.5 mL microcentrifuge tubes | Genesee Scientific | 24-281 | $25.90 | |
| Genie® Strips | Optigene | OP-0008-500 | $528.15 |
| Genie® II | Optigene | GEN2-01 | $10,000.00 |
SETTING UP THE GENIE II
30s
Plug In
Equipment
Genie® II
NAME
Heat Block and Reader
TYPE
Genie
BRAND
GEN2-01
SKU
15s
Set the block temperature to65 °C . Make sure temperatures are reached to set temperature before placing the assay tubes into heat-block for amplification.
15s
Setting up Workplace with Reagents
4m
Take Red Strips and Geen Strips out of freezer and thaw for 1-2 min at room temperature
2m
Set Blue Tubes in work area
10s
Set up biohazard disposal
10s
Take a swab
2m
Hand testee a sterile swab, packaged in 15 mL falcon tube, and instruct patient on use
30s
Testee to remove swab from 15 mL falcon tube
10s
Testee inserts swab into each nasal cavity, rotating for 10 seconds
30s
Testee returns swab to 15 mL falcon tube
30s
Testee hands tube to supervisor
10s
Preparing a sample batch (up to 8 samples different testees)
5m
Open sample swab with caution and transfer swab to a blue tube, which contains elution solution and mix/swirl the swab 4-5 times.
30s
Discard the swab in a biohazardous waste container
5s
Pipette 5 µL of eluted swab sample from the blue tube into a tube on the red strip, which contains SARS-CoV-2 master mix. Pipette up and down 2 times to mix assay components, then close the lid
20s
Pipette 5 µL of eluted swab sample from blue tube into a tube on the green strip, which contains the internal control master mix. NOTE: samples from each patient should be put in the same tube number on the red strip and green strip
20s
Repeat steps 4.1 through 4.4 for all samples in the batch
Open the
Equipment
Genie® II
NAME
Heat Block and Reader
TYPE
Genie
BRAND
GEN2-01
SKU
. Place the red batch into Block A and the green batch into Block B. Close the Genie II.
10s
Running a sample
30m
On the Genie II touch screen, select blocks A and B
10s
Run program called "FireLAMP"
10s
Wait for incubation to complete (~30 minutes for negative samples)
30m
Interpretation of results
1m
Touch results tab.
10s
Record results based on the "results column" column for block A and block B. Positive result in the block with the red strip (typically block A) indicate a positive result for COVID19. Positive results in the block with the green strip (typically block B) indicate that the sample was taken correctly.
10s
Record results
20s
Preparing for Next Block
5m
Discard all used blue tubes, red strips, and green strips in biohazard container
20s
Wait 5 minutes for Genie II to cool before starting analyis of a new batch.
5m