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

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-microliters 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 DQ alpha- and HIV-specific sequences. Treatment at 37 degrees C and 60% humidity for 7 days, storage for 12 weeks at 22 degrees C, and freeze-thawing twice had no adverse effect on PCR reactivity when compared with the results obtained with reference spots stored at -20 degrees 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.

Rapid DNA fingerprinting to control for specimen errors in HIV testing by the polymerase chain reaction.

Variable-number-tandem-repeats (VNTRs) are highly polymorphic and provide informative genetic markers for distinguishing between individuals. We have used PCR amplification of VNTR locus pMCT118 to identify mislabelled specimens submitted for HIV PCR testing. The method is rapid, can be applied to large numbers of samples and eliminates the need for radioactive probes. DNA samples (10 ng) are amplified for 25 cycles using fluorescence-labelled oligonucleotide primers (blue dye). An aliquot of the PCR product is then combined with an internal lane size standard (labelled with a red dye), electrophoresed through a 2% agarose gel on an automated fluorescence DNA fragment analyser and the size and quantity of the fragments determined automatically relative to the internal standard. Fifteen alleles, ranging in size from 398 tp 709 bp were readily identified in a random sampling of DNA from 63 unrelated HIV-infected patients. Fragment size was reproducible and corresponded to alleles containing from 16 to 35 repeats of a 16 bp unit. VNTR genotyping will prove useful for resolving discordant results due to specimen mix-up and ensuring that the correct samples have been analyzed.

Omeprazole 20 mg and 40 mg daily in the treatment of gastric ulcer: a pilot study in Filipino patients.

This study was an open evaluation of omeprazole, 20 mg or 40 mg once daily, for the treatment of gastric ulcer. Thirty-four Filipino patients, 22 with gastric (GU) and 12 with prepyloric (PPU) ulcers at least 5 mm in diameter were recruited. In terms of ulcer size and number of ulcers per patient, the 40 mg group had more severe ulcer disease. Twenty-two patients (15 GU and seven PPU) received 20 mg omeprazole and 12 patients (eight GU and four PPU) received 40 mg for 2-8 weeks, according to healing. Ulcers were assessed endoscopically every 2 weeks and a biopsy was performed (when unhealed) to exclude malignancy. Symptoms were recorded at each visit to the clinic and also in daily diary cards. Ulcer-healing rates were assessed for all patients who fulfilled protocol requirements at any one visit and were as follows at 2, 4, 6 and 8 weeks, respectively, for 20 mg: 50%, 70%, 85% and 95%; for 40 mg: 50%, 75%, 92% and 92%; and for all patients: 50%, 72%, 88%, and 94%. Results at 4, 6 and 8 weeks excluded two patients who were lost to follow-up. All patients who completed the study and whose ulcers were less than or equal to 20 mm in size (24) were healed as well as six of eight patients with ulcers greater than 20 mm. At entry, all but one patient reported symptoms. After only one dose of omeprazole, 50% of the patients on each dose no longer reported pain in their diary cards and after 4 weeks most patients were symptom-free.(ABSTRACT TRUNCATED AT 250 WORDS)