Methods
Study design and setting
We conducted a retrospective laboratory based cross-sectional study at the Livingstone University Teaching Hospital laboratory (LUTH). The LUTH laboratory is located in Livingstone town, southern part of Zambia. It is a main referral center for specialized testing for HIV VL and EID diagnosis servicing 13 districts in the province.
Sample size
120 samples (60 positives and 60 negatives) of less than or equal to 18 months were used (as per manufactures recommendation)
Quantitative:
HIV VL 154 samples
Open-Epi sample size calculator was used with 11.3% HIV prevalence in Zambia at 5% CI
EID DBS cards collected from different facilities that had been received at Livingstone laboratory for routine testing were conveniently sampled after being confirmed reactive and non-reactive on current platform COBAS Taqman (CAP/CTM) from infants of age £ 18 months. A sample size of 60 DBS cards with at least 2 spots (as per manufacturer recommendation) were retrieved from storage within a retention period of less than 18 months. For HIV VL, we conveniently sampled 154 HIV VL stored samples comprising of three levels; Target not detected, below titer and above titer. These were in EDTA containers collected within 24 hours and spotted on DBS cards and left to dry for 24 hours before processing on the Hologic Panther machine. We used results generated by COBAS 4800 to assign the levels.
Test methods
The COBAS Taqman (CAP/CTM) HIV-1 qual test is a qualitative nucleic acid amplification test for the detection of HIV type 1 RNA and proviral DNA in plasma, anti-coagulated fresh whole blood and DBS. The test is based on four major processes; Sample preparation and incubation, Sample preparation to isolate HIV-1 target nucleic acids-both processes utilizing the magnetic glass particle technology, Reverse transcription of the target RNA to generate complimentary DNA(cDNA) and Detection of cleaved dual labelled oligonucleotide detection probe specific to the target.
The COBAS4800 HIV-1 is based on fully automated sample preparation followed by PCR amplification and detection. The cobas® 4800 System consists of the cobas x 480 sample preparation instrument and the cobas z 480 real-time PCR analyzer. Real-time detection and discrimination of PCR products is accomplished by measuring the fluorescence of the released reporter dyes for the viral targets and RNA QS, respectively.
The Aptima HIV-1 Quant Dx assay involves three main steps, which all take place in a single tube on the Panther system: these include target capture, target amplification by transcription-mediated amplification (TMA), and detection of the amplification products (amplicon) by the fluorescent labeled probes (torches).
Statistical Methods
The collected data were entered in excel, thereafter exported to Stata version 15 for analysis. Chi square test was used for qualitative data to compare results from machine 1 COBAS Taqman (CAP/CTM) and machine 2 (Hologic panther). Pearson correlation test was to determine the relationship between quantitative data generated from COBAS4800 (machine 1) and Hologic panther (machine 2).
Reporting guidelines
We used the CLSI guidelines, EP15-A3 in our reporting and interpretation of the findings