Reactions contained 1 X WarmStart® Colorimetric LAMP Master Mix (M1800; NEB) supplemented with 1 x EvaGreen (Biotium), 0.02 U/μL Antarctic Thermolabile UDG (NEB), 700 μM dUTP (NEB), 1 X standard concentration LAMP primers and 40 mM guanidine chloride solution (Sigma G3272, pH adjusted to pH ~8). Reactions were prepared to final volume of 25 μl using nuclease free water, incubated at 65°C for 40 minutes on a StepOnePlus thermocycler (Applied Biosystems). The colour of finished reactions was recorded using an office flatbed scanner.
Warmstart colorimetric lamp master mix
Manufactured by New England Biolabs
Sourced in Canada
WarmStart® Colorimetric LAMP Master Mix is a ready-to-use solution for performing Loop-Mediated Isothermal Amplification (LAMP) reactions with colorimetric detection. It contains all the necessary components, including DNA polymerase, buffers, and a colorimetric dye, to enable rapid and sensitive nucleic acid amplification and detection.
Automatically generated - may contain errors
Lab products found in correlation
Betaine, by Merck Group (2 mentions)
Chemidoc mp imaging system, by Bio-Rad (2 mentions)
Guanidine chloride solution, by Merck Group (1 mentions)
Antarctic thermolabile udg, by New England Biolabs (1 mentions)
Steponeplus thermocycler, by Thermo Fisher Scientific (1 mentions)
Evagreen, by Biotium (1 mentions)
Geneamp pcr system 2700, by Thermo Fisher Scientific (1 mentions)
10 protocols using warmstart colorimetric lamp master mix
1
Colorimetric LAMP Detection with Guanidine
2
Rapid Shark Species Identification via LAMP
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LAMP was performed in a total volume of 10 μL containing 3 μL of nuclease-free water, 5 μL of WarmStart® Colorimetric LAMP Master Mix (NEB, UK), 1 μL of primers FIP (0.8 μM), BIP (0.8 μM), F3 (0.2 μM), B3 (0.2 μM), LF (0.4 μM) and specific primers for the three sharks (bigeye thresher, pelagic thresher and shortfin mako shark; 2) or Lambda (λ) LF and/or LR (0.4 μM), and 1 μL of DNA (approximately 5 ng/μL). Negative controls, containing no template DNA, were also prepared. LAMP primers for the positive control using λ DNA were obtained from Merck (Germany), and λ DNA was purchased from New England Biolabs (NEB, UK) (250 ng λ DNA, stored in 10 mM Tris HCl pH 7.5, 10 mM NaCl and 1 mM EDTA). For the positive control LAMP reaction, 5 ng/μL of λ DNA was used (more detail on controls can be found in SI 2).
The mixture was incubated at 65°C for 1 hour and then visualised on a 1.5% agarose gel, following electrophoresis, using a Biorad Gel Doc EQ system w/ Universal Hood II (UV transilluminator) (Bio-Rad Laboratories, USA). Successful amplification was indicated by a colour change (pink to yellow) and presence of bands on the gel.
The mixture was incubated at 65°C for 1 hour and then visualised on a 1.5% agarose gel, following electrophoresis, using a Biorad Gel Doc EQ system w/ Universal Hood II (UV transilluminator) (Bio-Rad Laboratories, USA). Successful amplification was indicated by a colour change (pink to yellow) and presence of bands on the gel.
3
Colorimetric LAMP for Rapid Detection
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All LAMP reactions were conducted in the WarmStart colorimetric LAMP master mix (NEB Canada, Whitby, ON, Canada). Each reaction was 25 μL in total and contained 1 μL template. Quantities of other components in each reaction followed the NEB’s instructions for the master mix. All reactions were conducted in individual 200-μL PCR tubes. The reaction program consisted of only one step in which the tubes were incubated at 65°C for 30 minutes. After the incubation, the reactions were checked visually and the results were recorded by photography.
4
CPNA-LAMP Diagnostic Assay Protocol
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CPNA-LAMP was performed using WarmStart® Colorimetric LAMP Master Mix (New England Biolabs, NEB), custom LAMP primers (IDT, Panagene), and 1M betaine (SigmaAldrich). LAMP master mix contains Bst polymerase, an enzyme derived from the large fragment of Bacillus stearothermophilus DNA Polymerase I. LAMP reactions were performed in nuclease free PCR tubes at a 25μL reaction volume. Primer concentrations were 0.2 μM F3, 0.2 μM B3, 1.6 μM FIP, 1.6 μM BIP, 0.8 μM R132H SALP, and 0.25 μM PNA. Reactions were incubated at 65°C for 45–60 minutes, then denatured at 95°C for 2 minutes. Amplification was confirmed via 4% agarose gel electrophoresis at 100 volts for 1 hour then imaged with a BioRad ChemiDoc MP™ Imaging System.
5
Colorimetric ASFV-LAMP Detection Protocol
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Colorimetric ASFV-LAMP reactions were performed under the following conditions: WarmStart® Colorimetric LAMP Master Mix (New England biolabs, Ipswitch, MA) 1X, FIP and BIP primers (0.8µM), LoopF and LoopB primers (0.4µM) and F3 and B3 primers (0.2µM). LAMP amplification was carried out at 65°C, either in a thermocycler (GeneAmp® PCR system 2700, Applied biosystems) or in a water bath, using pSLA DNA as template (5µL/reaction). Final reaction volume was set at 25µL. Amplification was carried out at 65°C for 30 minutes. Reactions were deemed as positive (bright yellow), suspect (pale pink) or negative (bright pink), according to the coloration of the mastermix at the end of the amplification.
6
LAMP-based MRSA detection on microfluidic chip
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Instead of eluting the surface-bound total RNA into nuclease-free water in the final step of the total RNA extraction on the chip, a mixture of WarmStart® Colorimetric LAMP Master Mix (New England BioLabs) and a customized primer set (1.6 µM for forward and backward inner primers, 0.2 µM for forward and backward outer primers, and 0.4 µM for forward and backward loop primers, Integrated DNA Technologies) was added to the chamber for on-chip amplification followed by colorimetric detection. The primer sequences for the nuc and mecA genes in MRSA are shown in Table S2
7
Optimized LNA-LAMP for Rapid Detection
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LNA-LAMP was performed using WarmStart® Colorimetric LAMP Master Mix (New England Biolabs, NEB), custom LAMP primers containing LNAs (IDT), and 1M betaine (Sigma-Aldrich). LAMP reactions were performed in nuclease-free PCR tubes at 25 or 50 µl reaction volumes. Primer concentrations were 0.2 µM F3, 0.2 µM B3, 1.6 µM FIP, 1.6 µM BIP, and 0.8 µM LNA-SALP. Reactions were optimized by incubation at temperatures ranging from 67 to 69°C for 35–45 min and then denatured at 95°C for 2 min. A final reaction temperature of 68°C was identified as the optimal temperature for the final selection of primers. Amplification was confirmed via 4% agarose gel (EtBr) electrophoresis at 100 volts for 1 h and then imaged with a BioRad ChemiDoc MP™ Imaging System.
8
Colorimetric LAMP Assay Protocol
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All LAMP reactions were conducted in the WarmStart colorimetric LAMP master mix (NEB Canada, Whitby, ON). Each reaction was 25 µL in total and contained 1 µL template. Quantities of other components in each reaction followed the NEB's instructions for the master mix. All reactions were conducted in individual 200-µL PCR tubes. The reaction program consisted of only one step in which the tubes were incubated at 65°C for 30 minutes. After the incubation, the reactions were checked visually and the results were recorded by photography.
9
LAMP-Based Colorimetric COVID-19 Detection
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The LAMP primer set Xt-CLS (Langlois et al. 2017) was used for confirmation of the qPCR test.
All LAMP reactions were conducted in the WarmStart colorimetric LAMP master mix (NEB Canada, Whitby, ON). Each reaction was 25 µL in total and contained 2 µL template. Quantities of other components in each reaction followed the NEB's instructions for the master mix. All reactions were conducted in individual 200-µL PCR tubes. The reaction program consisted of only one step in which the tubes were incubated at 65°C for 30 minutes. After the incubation, the reactions were checked visually and the results were recorded by photography.
All LAMP reactions were conducted in the WarmStart colorimetric LAMP master mix (NEB Canada, Whitby, ON). Each reaction was 25 µL in total and contained 2 µL template. Quantities of other components in each reaction followed the NEB's instructions for the master mix. All reactions were conducted in individual 200-µL PCR tubes. The reaction program consisted of only one step in which the tubes were incubated at 65°C for 30 minutes. After the incubation, the reactions were checked visually and the results were recorded by photography.
10
Colorimetric LAMP Assay for Detection
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All LAMP reactions were conducted in the WarmStart colorimetric LAMP master mix (NEB Canada, Whitby, ON). Each reaction was 25 µL in total volume containing 1 µL template.
Quantities of primers in each reaction followed the NEB's instructions for the master mix. All reactions were conducted in individual 200-µL PCR tubes. The reaction program consisted of only one step in which the tubes were incubated at 60°C for 50 minutes. After the incubation, the reactions were checked visually and the results were recorded by photography.
Quantities of primers in each reaction followed the NEB's instructions for the master mix. All reactions were conducted in individual 200-µL PCR tubes. The reaction program consisted of only one step in which the tubes were incubated at 60°C for 50 minutes. After the incubation, the reactions were checked visually and the results were recorded by photography.
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