Evaluation of influenza-specific humoral response by microbead array analysis.

RATIONALE: Quantitation and determination of antigen specificity of systemic and mucosal immune responses to influenza vaccination is beneficial for future vaccine development. Previous methods to acquire this information were costly, time consuming and sample exhaustive. The benefits of suspension microbead array (MBA) analysis are numerous. The multiplex capabilities of the system conserve time, money and sample, while generating statistically powerful data. OBJECTIVE: To demonstrate the use of the assay by comparing the humoral influenza-specific responses of two cohorts from two countries that differed in circulating influenza strains and rates of influenza vaccination. METHODS: Influenza hemagglutinin (HA) from different strains were coated on microbeads and incubated with serum samples to capture immunoglobulin (Ig) A(1) and IgG(1) host antibodies. RESULTS: Statistically significant differences in IgA(1) and IgG(1) exist between the serum samples from Winnipeg (Manitoba) donors and those from Kenyan (Africa) donors. Data were compared using Mann-Whitney nonparametric tests. The Winnipeg donors had higher mean fluorescence intensity values, with significant P values for anti-HA IgA(1) to A/Wyoming/3/2003 (P=0.044), A/Vietnam/1203/2004 (P=0.0179), A/New Caledonia/20/99 (P<0.0001) and B/Tokyo/53/99 (P=0.0002). No differences were seen between the groups in their response to B/Jilin/20/2003. The Winnipeg donors had higher mean fluorescence intensity values, with significant P values for anti-HA IgG(1) to A/Wyoming/3/2003 (P=0.0135), B/Tokyo/53/99 (P=0.006) and B/Jilin20/2003 (P=0.026). CONCLUSION: Influenza-specific IgA(1) and IgG(1) antibodies were successfully detected using MBA technology. A significant difference in antibody response was observed between Winnipeg and Kenyan donor serums. MBA analysis is a relatively quick and cost-effective method for serum antibody analysis. The potential to simultaneously assay small sample volumes for a multitude of antigens makes this method invaluable for future vaccine response monitoring.

Defining the ideal injection techniques when using 5-mm needles in children and adults.

OBJECTIVE: We aimed to establish the ideal injection techniques using 5-mm needles to reliably inject insulin into the subcutaneous fat in both children and adults and to quantify the associated pain and leakage of the test medium. RESEARCH DESIGN AND METHODS: A total of 259 subjects (122 children/adolescents and 137 adults) were injected with sterile air corresponding to 20 IU insulin (200 microl) with 32-G 5-mm needles at 90 degrees or 45 degrees , in the abdomen and thigh, and with or without a pinched skin fold. Injection depth was assessed via ultrasonography. Subjects rated pain on a visual analog scale. Test medium injections into the abdomen and thigh (0.2-0.6 ml) were also administered to assess injection leakage. RESULTS: Among children, 5.5% of injections were intramuscular (IM) and 0.5% were intradermal, while in adults, the incidence was 1.3 and 0.6%, respectively. The frequency of IM injections was greater in boys and negligible among adult women. Subcutaneous fat thickness was the primary predictor of the likelihood of IM injections (P < 0.001). A third of all patients reported experiencing no pain during insulin injection, with children/adolescents experiencing considerably more discomfort than adults. Some leakage of medium was observed, but was unrelated to injection volume and was generally minimal. CONCLUSIONS: 5-mm needles are reliably inserted into subcutaneous fat in both adults and children. These needles were associated with reduced pain and minimal leakage. We recommend an angled injection with a pinched skin fold for children, while in adults, the technique should be left to patient preference.

A common CD4 gene variant is associated with an increased risk of HIV-1 infection in Kenyan female commercial sex workers.

BACKGROUND: It has been predicted that CD4 C868T, a novel CD4 single-nucleotide polymorphism (SNP) that has been found to be highly prevalent among Africans, changes the tertiary structure of CD4, which may alter susceptibility to human immunodeficiency virus (HIV) infection. METHODS: Participants were from a Kenyan cohort and included 87 uninfected and 277 HIV-1-infected individuals. DNA sequencing was used to determine CD4 genotype. A2.01 cells expressing similar levels of either wild-type CD4 or CD4-Trp240 as well as peripheral blood mononuclear cells from uninfected donors were infected with HIV-1(IIIB) or a Kenyan primary HIV-1 isolate. HIV-1 p24 enzyme-linked immunosorbent assay was used to determine the outcome of infection. RESULTS: CD4 C868T was found to be significantly more prevalent among HIV-1-infected participants than among HIV-1-uninfected participants (P = .002), and C868T was associated with an increased incidence of HIV-1 infection as well (P = .005, log-rank test; P = .009, Wilcoxon test), with an odds ratio of 2.49 (P = .009). Both in vitro and ex vivo models demonstrated a significant association between CD4 C868T and susceptibility to HIV-1 infection (P < .001 and P = .003, respectively). CONCLUSION: Overall, the present study found a strong correlation between CD4 C868T and increased susceptibility to HIV-1 infection. Given the high prevalence of both HIV infection and CD4 C868T in African populations, the effect of this SNP on the epidemic in Africa could be dramatic.