Stability of dried blood spot specimens for detection of human immunodeficiency virus DNA by polymerase chain reaction

Abstract

Blood sampling on filter paper has many advantages for the detection of perinatal human immunodeficiency virus (HIV) infection by the polymerase chain reaction (PCR). However, if the method is to be widely used, an assessment of its performance under field conditions is required. To simulate conditions in the field, 50-I1 aliquots of whole blood containing low levels of HIV proviral DNA (4 to 1,024 copies per 100,000 nucleated cells) were spotted onto filter paper; dried; and subjected to heat, humidity, and prolonged storage at room temperature. After exposure, the DNA was recovered and amplified with primers to human leukocyte antigen DQoa-and HIV-specific sequences. Treatment at 37°C and 60%o humidity for 7 days, storage for 12 weeks at 22°C, and freeze-thawing twice had no adverse effect on PCR reactivity when compared with the results obtained with reference spots stored at-20°C. The lower limits of HIV detection in all tests ranged from 4 to 16 HIV copies per 100,000 cells. Fixation in 70% ethanol improved the amplification of low levels of HIV DNA and reduced biohazard risks. These findings suggest that dried blood spots will provide a powerful new resource for testing for HIV by PCR, especially in remote areas where refrigeration and immediate sample processing are unavailable.

Single Lysis‐Salting Out Method of Genomic DNA Extraction From Dried Blood Spots

“…The first step in SLSO method is the addition of methanol to DBS punches to prevent diffusion of natural inhibitors such as heme and its degradation products from the filter paper matrix, which might considerably pose technical difficulties while performing PCR amplifications . The SLSO method was carried out in two phases, phase I includes one‐step lysis to degrade cellular contents using concentrated lysis buffer, which results in improved lysis with minimal sample loss and contamination.…”

Detection of HIV-1 infection in dried blood spots from a 12-year-old ABO bedside test card

“…Several studies have been reported on the use of spots of dried blood on filter paper for HIV testing (for example, of newborns or persons living in remote areas) by means of PCR [7][8][9][10]. Other reports describe the successful use of such blood spots in newborns for the detection of maternal HIV [11,12] or HTLV-I [13] antibodies for epidemiological purposes.…”

Exposed HIV infants who had their DBS sample collected, tested and received result in past 12 months (January-December 2013) at Specialist Hospital Yola, Nigeria

“…HIV attacks the human immune system, the T helper cell predominantly CD4 + T cells, macrophages, and dendritic cells. [3] HIV infection depletes CD4 + T cells to a low level by mechanisms such as apoptosis of cells, [4] direct killing of infected cells, and killing by CD8 cytotoxic lymphocytes. [5] When CD4 + T cell numbers decline below a critical level, cellmediated immunity is lost, and the body becomes progressively more susceptible to opportunistic infections.…”

“…An estimated prevalence of HIV infection in adults between the ages of 15-49 is 7% and children less than 5 years is 1.7% in Uganda [2]. With early case detection of children infected with HIV with the aid of early infant diagnosis (EID) has helped in the early initiation of antiretroviral therapy, thereby reducing morbidity and mortality [3,4]. Currently, EID testing in Nigeria is done by polymerase chain reaction (PCR) analysis of HIV DNA samples in centralized laboratories.…”