Internally Controlled, Generic Real-Time PCR for Quantification and Multiplex Real-Time PCR with Serotype-Specific Probes for Serotyping of Dengue Virus Infections

Table 3

Dynamic range of generic real-time PCR^a.

DENV in vitro RNA copies in extraction	DENV in vitro RNA copies in RT	DENV in vitro RNA copies in PCR^b	IC copies in PCR^b	Number positives DENV in vitro RNA (%; mean Cq-value: SD)	Number positives IC (%; mean Cq-value: S.D)
1.25×10^7	10^7	10^6	500	12/12 (100%; 26.59; 0.54)	12/12 (100%; 35.82; 1.08)
1.25×10^6	10^6	10^5	500	12/12 (100%; 30.59; 0.46)	12/12 (100%; 32.67; 0.79)
1.25×10^5	10^5	10^4	500	12/12 (100%; 34.35; 1.09)	12/12 (100%; 32.97; 0.74)
1.25×10^4	10^4	10^3	500	8/12 (66.7%; 37.22; 0.73)	12/12 (100%; 33.73; 0.63)
1.25×10^3	10^3	10^2	500	1/12 (8%; 40.71)	12/12 (100%; 32.24; 0.60)
0	0	0	0	0/12	0/12

^aThe dynamic range of the generic PCR was determined on 10-fold dilution series of DENV in vitro RNA in the presence of a fixed amount of IC spiked into lysis buffer containing DENV negative human serum as background and for 12 replicates per dilution.
^bNumber of copies cDNA/PCR has been calculated as 1/12.5 of the extracted RNA. assuming 100% efficiency in extraction, reverse transcription (RT), and PCR.
SD: standard deviation.