Superiority of MALDI-TOF Mass Spectrometry over Real-Time PCR for SARS-CoV-2 RNA Detection
Abstract
:1. Introduction
2. Materials and Methods
2.1. Sample Collecton, Storage and Workflow
2.2. Real–Time PCR
2.3. Mass Spectrometry-Based Assay
3. Results
4. Discussion
5. Conclusions
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
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Reagent | Per Reaction (µL) |
---|---|
RT-PCR | |
10×PCR Buffer | 0.500 |
MgCl2, 25mM | 0.400 |
dNTP Mix, 25mM | 0.100 |
UNG | 0.050 |
RNase Inhibitor | 0.125 |
PCR Enzyme | 0.200 |
MMLV Enzyme | 0.125 |
SARS-CoV-2 PCR Primers | 0.500 |
Sample RNA | 3 |
SAP Cocktail | |
HPLC-grade water | 1.53 |
SAP Buffer | 0.17 |
Shrimp Alkaline Phosphatase (SAP) | 0.3 |
Extension Cocktail | |
HPLC-grade water | 0.62 |
iPLEX Buffer Plus, GPR | 0.20 |
iPLEX Termination Mix | 0.20 |
iPLEX Pro Enzyme | 0.04 |
SARS-CoV-2 Extend Primers | 0.94 |
SARS-CoV-2 Targets | MS2 | QC Status | Judgement |
---|---|---|---|
≥2 SARS-CoV-2 target detected | Detected | Passed | Positive |
<2 SARS-CoV-2 target detected | Detected | Passed | Suspected |
0–5 SARS-CoV-2 target detected | Not detected | Failed | Invalid |
Sample No | Official Report by RT-PCR a | Results of RT-PCR b | Results of MassARRAY®,c | MassARRAY® Judgment | |||||
---|---|---|---|---|---|---|---|---|---|
ORF1ab | N | ORF1ab | ORF1 | N1 | N2 | N3 | |||
1 | S | 35.92 | ND | ND | P | P | P | P | P |
2 | S | ND | 38.37 | ND | P | P | ND | P | P |
3 | S | ND | ND | ND | P | ND | ND | ND | S |
4 | S | ND | ND | P | ND | ND | P | ND | P |
5 | S | ND | 36.24 | ND | P | P | ND | P | P |
6 | S | ND | 36.85 | ND | P | P | P | P | P |
7 | S | ND | 34.26 | P | ND | ND | P | ND | P |
8 | S | ND | 37.20 | ND | P | P | ND | P | P |
9 | S | ND | ND | ND | P | ND | ND | ND | S |
10 | S | ND | 35.96 | ND | ND | P | P | ND | P |
11 | S | 34.72 | ND | P | P | P | ND | P | P |
12 | S | 36.20 | ND | P | P | P | P | P | P |
13 | S | 35.44 | ND | ND | ND | P | P | P | P |
14 | S | ND | 35.35 | P | P | P | P | P | P |
15 | S | ND | 35.48 | P | P | P | P | P | P |
16 | N | ND | ND | ND | ND | ND | ND | ND | N |
17 | N | ND | ND | ND | P | ND | ND | ND | S |
18 | N | ND | ND | ND | ND | ND | ND | ND | N |
19 | N | ND | ND | ND | ND | ND | ND | ND | N |
20 | N | ND | ND | ND | ND | ND | ND | ND | N |
21 | N | ND | ND | ND | ND | ND | ND | ND | N |
22 | N | ND | ND | ND | ND | P | ND | P | P |
23 | N | ND | ND | ND | ND | ND | ND | ND | N |
24 | N | ND | ND | ND | P | ND | ND | ND | S |
25 | N | ND | ND | ND | ND | ND | ND | ND | N |
26 | N | ND | ND | ND | ND | ND | ND | ND | N |
27 | N | ND | ND | ND | ND | ND | ND | ND | N |
28 | N | ND | ND | ND | ND | ND | ND | ND | N |
29 | N | ND | ND | ND | P | P | ND | P | P |
30 | N | ND | ND | ND | ND | ND | ND | ND | N |
31 | N | ND | ND | ND | ND | ND | ND | ND | N |
32 | N | ND | ND | ND | ND | ND | ND | ND | N |
33 | N | ND | ND | ND | ND | ND | ND | ND | N |
34 | N | ND | ND | ND | ND | ND | ND | ND | N |
35 | N | ND | ND | ND | ND | ND | ND | ND | N |
Technique | RT-PCR | MassARRAY® System |
---|---|---|
Targets | ORF1ab, N | ORF1ab, ORF1, N1, N2, N3 |
RNA volume | 5 µL | 3 µL |
Time needed * | 4–6 h | 8 h |
Cost | ~10 EUR/sample | ~10 EUR/sample |
Report method | Qualitative | Qualitative |
Advantages | Short time of analyses; specific, confirms active cases; useful in clinical decision-making | Highly sensitive, accurate, high-throughput, easy data analysis, cost-effective, simple partially automated workflow, runs up to eight 96-well plates per day, may process two more plates overnight. |
Disadvantages | Unable to simultaneously detect virus and its mutations, less accurate in low viral load samples | Multiple preparation steps, requires specialized equipment |
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Rybicka, M.; Miłosz, E.; Bielawski, K.P. Superiority of MALDI-TOF Mass Spectrometry over Real-Time PCR for SARS-CoV-2 RNA Detection. Viruses 2021, 13, 730. https://doi.org/10.3390/v13050730
Rybicka M, Miłosz E, Bielawski KP. Superiority of MALDI-TOF Mass Spectrometry over Real-Time PCR for SARS-CoV-2 RNA Detection. Viruses. 2021; 13(5):730. https://doi.org/10.3390/v13050730
Chicago/Turabian StyleRybicka, Magda, Ewa Miłosz, and Krzysztof Piotr Bielawski. 2021. "Superiority of MALDI-TOF Mass Spectrometry over Real-Time PCR for SARS-CoV-2 RNA Detection" Viruses 13, no. 5: 730. https://doi.org/10.3390/v13050730
APA StyleRybicka, M., Miłosz, E., & Bielawski, K. P. (2021). Superiority of MALDI-TOF Mass Spectrometry over Real-Time PCR for SARS-CoV-2 RNA Detection. Viruses, 13(5), 730. https://doi.org/10.3390/v13050730