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EID - Current tools and prospects of point of care technology

EID - Current tools and prospects of point of care technology. Susan A. Fiscus, Ph.D. University of North Carolina at Chapel Hill. Disclosures. Honoraria: Gen-Probe, Roche, Abbott Free kits: Roche, Gen-Probe, Perkin-Elmer, Cavidi, Siemans, Abbott, Inverness, IQuum, ImmunoDiagnostics. Outline.

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EID - Current tools and prospects of point of care technology

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  1. EID - Current tools and prospects of point of care technology Susan A. Fiscus, Ph.D. University of North Carolina at Chapel Hill

  2. Disclosures • Honoraria: Gen-Probe, Roche, Abbott • Free kits: Roche, Gen-Probe, Perkin-Elmer, Cavidi, Siemans, Abbott, Inverness, IQuum, ImmunoDiagnostics

  3. Outline • Desirable qualities for POC assays • Assays used for EID • HIV DNA • HIV RNA • P24 Antigen • Considerations when selecting an EID assay • Key points • Steps to move forward

  4. Desirable Qualities of a POC Test • Inexpensive (< $USD 5 /test) • Rapid (< 1 hour) • Simple • Equipment – battery operated, few moving parts, small footprint • Technique – minimal training required • Sensitive (how sensitive? > 95%?) • Specific (how specific? > 98%?) • Robust - No cold chain requirement • Commercially available/CE marked-FRA cleared

  5. Desirable Qualities of a POC Test • “Cheap, fast, or accurate. Pick 2” (Dr. Bill Rodriguez, Harvard Univ, Nov 16, 2009)

  6. HIV DNA Assays • Roche AMPLICOR HIV DNA assay, v 1.5 is the gold standard – either using whole blood pellets or Dried Blood Spots (DBS) • Has been successfully introduced and implemented in many countries around the world

  7. POC HIV DNA Assays • CIGHT, Kelso, Northwestern Univ- LoD 5 cp/reaction (Jangam, 2010, CROI); not yet ready for field testing and on hold while work focuses on a POC p24 test • Micronics – Real Time PCR (Tim Granade, CDC; CROI 2010) • BioHelix – isothermal lateral flow – 2 hr TAT (Jeanne Jordan, GWU) • Both Micronics and BioHelix seem to be more in the proof of concept stage and don’t yet seem ready for field testing.

  8. HIV RNA Assays • Have been used as alternatives to HIV DNA testing • Quantitative HIV RNA assays may not be as sensitive when infants are being prophylaxed or if mothers are receiving ARVs and the child is breast-feeding • Qualitative Gen-Probe Aptima • Very sensitive and specific (Kerr, 2009; Stevens, 2009) • Works well with DBS • Only HIV RNA assay FDA approved for diagnosis (though approval is for plasma or serum, not DBS) • Being used by the State of New York for EID

  9. Commercially Available FDA Cleared HIV-1 RNA Assays

  10. POC RNA Assays • IQuum – realtime PCR, LoD – ~100 cp/mL, 1 hr, 200 uL plasma • Inverness – microarray, realtime detection,10 uL whole blood • Helen Lee – semiquantitative dipstick with 200 cp/mL cutoff (Lee, JID 2010) • Advanced Liquid Logic - based on digital microfluidics • Wave 80 – assay based on bDNA assay

  11. LIAT™ Quantitative POC HIV Assay • 200 uL plasma sample input (haven’t tested whole blood yet) • Limit of detection - 78 copies/mL of ARNA detected in 60 min • Each cartridge has an internal control • Dynamic range 100 to 10 million cp/mL in 60 min • Detects HIV-1 Groups M and O and HIV-2 viruses • Comparative data with 30 clinical specimens: • Roche COBAS - 88.4% correlation coefficient • Siemens Versant – 92% correlation coefficient

  12. LIAT • 92% correlation with Abbott m2000 with 75 clinical specimens (clades A, B, C, and D) • Training took 10 minutes • Easy to use • Assay takes 60 minutes Fiscus, unpublished data 2010

  13. IMI’s CLONDIAG HIV Viral Load • Point-of-Care Test • Allows determination of HIV load in fingerstick, whole blood, or plasma. • Multiple HIV-1 and HIV-2 targets are detected simultaneously by a proprietary microarray real time detection method. • The test includes internal controls • The sample is applied directly onto the test cartridge • The cartridge is processed by a compact, battery driven instrument

  14. IMI IMI negative positive COBAS plasma 73 (28 %) 56 (22 %) negative COBAS plasma 8 (3 %) 21( 8 %) <40 cp/ml COBAS plasma 6 (2%) 94 (36%) positive donors receiving HAART therapy naïve donors blood viral load equals plasma viral load ——— IMI CLONDIAG HIV VL Test Data generated on 1 ml of EDTA Plasma (COBAS Ampliprep/Taqman) versus 10 µl of Whole Blood (IMI’s prototype assay) Percentage of samples with detectable viral load: COBAS (1 ml plasma) 50 % IMI VL (10 µl blood) 66 % all samples are from HIV-positive donors specificity of both assays =100% (32 HIV-negative donors)

  15. SAMBA HIV-1 POC Test Lee, et al 2010. JID 201 Suppl 1:S65-72

  16. SAMBA HIV-1 POC Test Comparison of the Simple Amplification-Based Assay (SAMBA) HIV-1 Test with Commercially Available HIV-1 Nucleic Acid Tests From Lee, et al 2010. JID 201 Suppl 1:S65-72

  17. Total Nucleic Acid Assays • Roche Taqman - CAP/CTM HIV-1 Qual is being introduced (Stevens, et al, JCM 2008) • Works on whole blood and DBS – 100% sensitive/99.7% specific • Abbott is developing a total nucleic acid assay as well • Both require expensive new instrumentation – Roche Taqman or Abbott m2000 • Limited information about the performance of these assays in more resource constrained settings • Probably suitable for large centralized labs

  18. HIV-1 p24 Antigen Tests • The ultrasensitive, heat dissociated p24 antigen assay has been shown to work well for EID • With both plasma • sensitivity - 91-100% • specificity - 95-100% • N= 2314 samples from 9 publications • And DBS • Sensitivity – 98-100% • Specificity – 100% • N=1328 from 3 publications

  19. Point of Care p24 Antigen Tests • Inverness Determine Combination Ab/p24 Ag • ImmunoDiagnostics • mBio Diagnostics • Northwestern –Abbott partnership - David Kelso

  20. p24 Antigen Rapid Test forDiagnosis of Acute Pediatric HIV Infection Plasma volume:25mL Immune Disruption:90oC heat shock Assay Steps:1. Add 25mL plasma to 75mL buffer2. Heat in water bath for 4min 3. Insert test strip &read after 20 min. Assay Sensitivity:50pg/mL or 40,000 RNA copies/mL p24 Rapid Test Strip

  21. Results from pre-clinical trials in Cape Town 394 infant samples tested at NHLS Virology Lab, Groote Schuur Hospital, Cape Town, South Africa 86% of samples were from infants under 6 months of age,53% from infants under 2 months of age Reference Assay: Total Nucleic Acid PCR (Roche Ampliprep/COBAS Taqman HIV-1) p24 Assay Sensitivity: 23/24 = 95.8% (95% CI 80-99%) p24 Assay Specificity: 363/365 = 99.4% (95% CI 98-100%) 5 samples gelled (1.3%) giving invalid results

  22. Point-Of-Care p24 Antigen Rapid TestUnder Development Assay Procedure 1. Separate plasma Whole blood volume: 80mL Immune Disruption: Heat shock Total Assay Duration: 35 min. Consumables: Plasma separator, reaction tube, reaction buffer, rapid test strip Processor: Battery operated Cost per Assay: $1-2 per test Ready early 2012? 2. Pretreat sample in processor 3. Insert rapid test strip and read results

  23. “Cheap, fast, or accurate. Pick 2”

  24. Factors to be Considered When Selecting an EID Assay • Performance characteristics • Sensitivity and specificity • Specimen type and volume • HIV subtype(s) in the population • Technical and support issues • Volume and throughput – 1-2 or 1000/day • Equipment footprint • Printable results • Training requirements • Acceptance by MOH and clinicians • External and internal quality assurance

  25. Centralized Testing using DBS Can be implemented now Better control on training, supply logistics, internal and external QA Potential for high through-put - - - - - - - - - - - - - - - - - - - - - - - Huge backlog of DBS in some countries with long turn around times Delays and problems in returning results Point of Care Results ready in an hr or less Possibly fewer problems with mislabeling Able to confirm positive test results immediately - - - - - - - - - - - - - - - - - - - - Potential problems with training, competency, logistics Not yet ready for prime time Centralized vs POC Testing

  26. Key Points • POC assays should be inexpensive, rapid, simple, sensitive, specific, and robust • Promising POC assays today include: IQuum’s LIAT, SAMBA, CIGHT’s p24, and possibly Clondiag’s • Timeline for field testing and implementation: • CIGHT p24 – Late 2011-early 2012 • IQuum - ???? • Inverness - ???? • SAMBA - ?????

  27. Steps to move forward • Continue lab validation of new POC tests • Field test new assays under controlled conditions • Expand usage and evaluate the effects of POC on key operational parameters: • % of infants tested • % of infants who receive their results • % of infected infants who access care • % of infected infants who die or are hospitalized before age 2 years

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