Collection of blood for monitoring of HIV viral load may be achievable using dried blood spots without sacrificing accuracy of measurement, according to a study published in the December 23rd 2003 issue of The Lancet.
Currently, in most African countries, laboratories that perform HIV diagnostics are located at only one or two hospitals in the largest cities. Unless patients are able to travel to the city, a clinic providing HIV therapy for persons in a rural setting must collect blood samples and have them transported to the central lab for processing. The results must then be sent back to the clinic where the medical staff can use them to make decisions about starting or changing an individual’s antiretroviral regimen.
The need to transport blood sample to the lab presents several problems. First, collection at the site requires that venous blood be taken from the individual by a competent phlebotomist equipped with a supply of needles, collection tubes and shipping containers. If plasma is to be sent, then the blood must be centrifuged at the site. Samples must be refrigerated and shipped in dry ice. During the trip to the lab, high temperatures and uncertain delays can affect blood samples. Delays in obtaining results means delay in initiating the proper treatment.
To minimise these variables, dried blood spots (DBS) and dried plasma spots (DPS) placed on filter paper have been investigated as alternative methods for transporting blood samples used for HIV viral load tests. Liquid blood samples are still required for performing CD4 counts by flow cytometry. While both DBS and DPS may be more durable in transit than liquid samples, DPS would still require collecting venous blood and centrifugation at the remote site. Dried whole-blood spots, then, offers the significant advantages of a minimally invasive collection technique (finger stick), without the need for specialised equipment, in addition to durability during transport.
Laboratory-based evaluations comparing dried plasma spots (DPS) to liquid plasma have previously demonstrated a good correlation of viral load results in African patients. A recent field evaluation of dried whole-blood spots published in The Lancet now provides a stepping-stone towards full validation for using DBS for viral load measurements in Africa and other resource-limited settings. The study was a collaboration between Dr. Peter Mwaba, of the University Teaching Hospital in Lusaka, Zambia, with the University of Ottawa in Ontario, Canada, and the Medical Research Council in London.
The researchers collected blood samples from 51 HIV-positive individuals attending the hospital clinic in Lusaka. Five whole blood spots obtained by finger stick were collected onto a filter paper Gutherie card. Venous blood was also collected and the extracted plasma similarly spotted onto a separate Gutherie card. In order to mimic the likely conditions in which samples could be stored and transported, the cards were labeled and placed into a bond paper ziplock envelope with a silica desiccant, then inserted into a shipping envelope. These envelopes were stored at room temperature for six weeks then transported from Zambia to Ottawa for HIV RNA analysis with a nucleic acid amplification (NASBA) technique adapted for dried sample spots. The results obtained with each collection technique were plotted against the mean of the samples and the differences correlated with the mean using the Spearman correlation coefficient.
Overall, the differences between DBS and DPS values obtained were very small, with values below 5.0 log copies of HIV RNA showing the least divergence. This study, by building on previous work that correlated DPS with liquid plasma, is an important step towards validating the routine use of DBS as a “field-friendly” method for collecting viral load samples in remote settings in Africa and elsewhere. Continuing research on DBS is needed to better understand the effects of variables such as drying time, humidity and temperature on RNA stability. The authors recommend that multi-country collaborative work be pursued “to refine the technique and show its practical usefulness in serving HIV-infected populations in remote regions.” They also point to the potential value of DBS for studying subtype and genotype prevalence as well as for the surveillance of drug resistance and superinfection, as treatment becomes more widely available in Africa.
While the deployment of additional and cheaper diagnostics technology and the simplification of sample transport that DSB allows will certainly make laboratory monitoring of antiretroviral therapy more accessible, some see this as only a bridge to a breakthrough that brings both viral load and CD4 measurements to the field level. Gregg Gonsalves, an advocate working towards the development of inexpensive, single-use dipstick assays, said, “dried blood spots are a simpler collection technique, but they still depend on high tech assays. This won’t help with diagnosis at the point-of-care where the treating staff need results in real-time. We need both simpler collection techniques and simpler assays.”
Further information on this website
Monitoring where resources are limited - a guide to lowering the costs of diagnostics and issues in the use of diagnostic tests in resource-limited settings
New CD4 counting method may lead to cheaper tests in developing world - news story, October 2003
Clinton Foundation secures cut price CD4 and viral load tests for resource limited countries - news story, January 15 2004
Mwaba P et al. Whole blood versus plasma spots for measurement of HIV-1 viral load in HIV-infected African patients. The Lancet 362: 2067, 2003.