TABLE 1.
Summary of participating laboratories responses to the technical questionnaire
| Analysis yr | Extraction method (%) | Target combination (%) | Real-time apparatus type (%)b | No. of cycles (%) | Probe type (%) | Amplification mixture (%) | Use of extraction (%)/amplification control(s) (%) |
|---|---|---|---|---|---|---|---|
| 2010 (n = 32a) | |||||||
| No. of answers | 28 | 31 | 30 | 29 | 30 | 30 | 29 |
| Commercial kit (50) | IS481/IS1001 (47) | LightCycler (Roche) (37) | 40 (55) | Hydrolysis (73) | Commercial kit (33) | 76/100 | |
| Automated system (36) | IS481 (44) | SmartCycler (Cepheid) (26) | 45 (41) | Hybridization (17) | PCR master mix completed with oligosc (67) | ||
| In-house method (14) | IS481/IS1001/ptxA or ptxA-Pr (6) | ABI Prism (Applied Biosystems) (20) | 35 (4) | SYBR Green (7) | |||
| IS481/ptxA (3) | Rotor-Gene (10) | Beacon (3) | |||||
| 2012 (n = 34a) | |||||||
| No. of answers | 34 | 34 | 34 | 34 | 34 | 34 | 34 |
| Commercial kit (52) | IS481/IS1001 (71) | LightCycler (Roche) (40) | 45 (59) | Hydrolysis (79) | Commercial kit (62) | 84/97 | |
| Automated system (38) | IS481 (23) | SmartCycler (Cepheid) (35) | 40 (26) | Hybridization (15) | PCR master mix completed with oligosc (38) | ||
| In-house method (13) | IS481/ptxA (3) | ABI Prism (Applied Biosystems) (12) | 50 (15) | SYBR Green (3) | |||
| IS481/IS1001/ptxA-Pr/recA-h (3) | Rotor-Gene (9) | Beacon (3) |
a
Participants using RT-PCR methodology are presented. The only laboratory using endpoint PCR method (Genoquick Bordetella Hain LifeScience kit; IS481-IS1001; Perkin Elmer apparatus) is not included in the table.
b
LightCycler 1/2/480; SmartCycler I/II; ABI Prism 7000/7500/7900; Rotor-Gene 6000.
c
Participants used RT-PCR mixture from different suppliers (Applied Biosystems, Roche, Eurogentec) completed with primers and probes from different suppliers.