Targeted Deposition of Antibodies on a Multiplex CMOS Microarray and Optimization of a Sensitive Immunoassay Using Electrochemical Detection

The CombiMatrix ElectraSense microarray is a highly multiplex, complementary metal oxide semiconductor with 12,544 electrodes that are individually addressable. This platform is commercially available as a custom DNA microarray; and, in this configuration, it has also been used to tether antibodies (Abs) specifically on electrodes using complementary DNA sequences conjugated to the Abs.An empirical method is described for developing and optimizing immunoassays on the CombiMatrix ElectraSense microarray based upon targeted deposition of polypyrrole (Ppy) and capture Ab. This process was automated using instrumentation that can selectively apply a potential or current to individual electrodes and also measure current generated at the electrodes by an enzyme-enhanced electrochemical (ECD) reaction. By designating groups of electrodes on the array for different Ppy deposition conditions, we determined that the sensitivity and specificity of a sandwich immunoassay for staphylococcal enterotoxin B (SEB) is influenced by the application of different voltages or currents and the application time. The sandwich immunoassay used a capture Ab adsorbed to the Ppy and a reporter Ab labeled for fluorescence detection or ECD, and results from these methods of detection were different.Using Ppy deposition conditions for optimum results, the lower limit of detection for SEB using the ECD assay was between 0.003 and 0.01 pg/ml, which represents an order of magnitude improvement over a conventional enzyme-linked immunosorbant assay. In the absence of understanding the variables and complexities that affect assay performance, this highly multiplexed electrode array provided a rapid, high throughput, and empirical approach for developing a sensitive immunoassay

Porphyrin-modified antimicrobial peptide indicators for detection of bacteria

This study demonstrates the potential of porphyrin modified antimicrobial peptides for indication of bacterial targets on the basis of changes in the spectrophotometric characteristics of the construct. Detection is a result of changes in the structure of the antimicrobial peptide upon target binding. Those constructs comprised of peptides that offer little or no change in conformation upon interaction with bacterial cells demonstrated negligible changes in absorbance and fluorescence when challenged using Escherichia coli or Bacillus cereus. CD analysis confirms the presence/absence of conformational changes in the porphyrin-peptide constructs. Differing spectrophotometric responses were observed for constructs utilizing different peptides. The incorporation of metals into the porphyrin component of the constructs was shown to alter their spectrophotometric characteristics as well as the resulting absorbance and fluorescence changes noted upon interaction with a target. The described constructs offer the potential to enable a new type of biosensing approach in which the porphyrin-peptide indicators offer both target recognition and optical transduction, requiring no additional reagents

Identification of \u3ci\u3ebla\u3c/i\u3e\u3csub\u3eOXA-51-like\u3c/sub\u3e, \u3ci\u3ebla\u3c/i\u3e\u3csub\u3eOXA-58\u3c/sub\u3e, \u3ci\u3ebla\u3c/i\u3e\u3csub\u3eDIM-1\u3c/sub\u3e, and \u3ci\u3ebla\u3c/i\u3e\u3csub\u3eVIM\u3c/sub\u3e Carbapenemase Genes in Hospital \u3ci\u3eEnterobacteriaceae\u3c/i\u3e Isolates from Sierra Leone

We describe the results of a molecular epidemiological survey of 15 carbapenemase-encoding genes from a recent collection of clinical isolates from Mercy Hospital in Bo, Sierra Leone. The most salient findings revealed that (i) 60% of the isolates harbored multiple carbapenemase genes; (ii) the blaDIM-1 gene, which has previously only been reported in The Netherlands, is also circulating in this environment; and (iii) blaOXA-51-like and blaOXA-58 genes, which were thought to reside exclusively in Acinetobacter species, can also be found in members of the Enterobacteriaceae

Surveillance of Vector-Borne Infections (Chikungunya, Dengue, and Malaria) in Bo, Sierra Leone, 2012–2013

Malaria remains a significant cause of morbidity and mortality in West Africa, but the contribution of other vector-borne infections (VBIs) to the burden of disease has been understudied. We used rapid diagnostic tests (RDTs) for three VBIs to test blood samples from 1,795 febrile residents of Bo City, Sierra Leone, over a 1-year period in 2012–2013. In total, 24% of the tests were positive for malaria, fewer than 5% were positive for markers of dengue virus infection, and 39% were positive for IgM directed against chikungunya virus (CHIKV) or a related alphavirus. In total, more than half (55%) of these febrile individuals tested positive for at least one of the three VBIs, which highlights the very high burden of vector-borne diseases in this population. The prevalence of positives on the Chikungunya IgM and dengue tests did not vary significantly with age (P > 0.36), but higher rates of malaria were observed in children < 15 years of age (P < 0.001). Positive results on the Chikungunya IgM RDTs were moderately correlated with rainfall (r 2 = 0.599). Based on the high prevalence of positive results on the Chikungunya IgM RDTs from individuals Bo and its environs, there is a need to examine whether an ecological shift toward a greater burden from CHIKV or related alphaviruses is occurring in other parts of Sierra Leone or the West African region

Prevalence of markers of HIV infection among febrile adults and children in Bo, Sierra Leone, 2012-2013.

The goal of this study was to examine the prevalence of HIV among febrile patients seeking care in Mercy Hospital, Bo, Sierra Leone, in 2012-2013. A total of 1207 febrile persons were tested for HIV with Determine™ and SD Bioline rapid diagnostic tests kits that detect the presence of HIV antibodies and HIV p24 antigens. The overall prevalence of HIV among the tested patients was 8.9%, which is considerably higher than the < 2% prevalence of HIV reported previously in the general population. While these results are not sufficient to prove a causal relationship, the obtained data imply that HIV positive individuals may be more likely to suffer from febrile infectious diseases than individuals without HIV infection. Increasing the availability and use of HIV testing services will allow antiretroviral therapy to be accessed in a timely manner and improve health status among people living with HIV