Parallel detection approaches are of interest to many researchers interested in identifying multiple water and foodborne pathogens simultaneously. Availability and cost-effectiveness are two key factors determining the usefulness of such approaches for laboratories with limited resources. In this study, we developed and validated a high-density microarray for simultaneous screening of 14 bacterial pathogens using an approach that employs gold labeling with silver enhancement (GLS) protocol. In total, 8887 probes (50-mer) were designed using an in-house database of virulence and marker genes (VMGs), and synthesized in quadruplicate on glass slides using an in-situ synthesis technology. Target VMG amplicons were obtained using multiplex polymerase chain reaction (PCR), labeled with biotin, and hybridized to the microarray. The signals generated after gold deposition and silver enhancement, were quantified using a flatbed scanner having 2-μm resolution. Data analysis indicated that reliable presence/absence calls could be made, if: i) over four probes were used per gene, ii) the signal-to-noise ratio (SNR) cutoff was greater than or equal to two, and iii) the positive fraction (PF), i.e., number of probes with SNR≥2 for a given VMG was greater than 0.75. Hybridization of the array with blind samples resulted in 100% correct calls, and no false positive. Because amplicons were obtained by multiplex PCR, sensitivity of this method is similar to PCR. This assay is an inexpensive and reliable technique for high throughput screening of multiple pathogens.
Journal Of Microbiological Methods – Elsevier
Published: Aug 1, 2017
It’s your single place to instantly
discover and read the research
that matters to you.
Enjoy affordable access to
over 18 million articles from more than
15,000 peer-reviewed journals.
All for just $49/month
Query the DeepDyve database, plus search all of PubMed and Google Scholar seamlessly
Save any article or search result from DeepDyve, PubMed, and Google Scholar... all in one place.
Get unlimited, online access to over 18 million full-text articles from more than 15,000 scientific journals.
Read from thousands of the leading scholarly journals from SpringerNature, Elsevier, Wiley-Blackwell, Oxford University Press and more.
All the latest content is available, no embargo periods.
“Hi guys, I cannot tell you how much I love this resource. Incredible. I really believe you've hit the nail on the head with this site in regards to solving the research-purchase issue.”Daniel C.
“Whoa! It’s like Spotify but for academic articles.”@Phil_Robichaud
“I must say, @deepdyve is a fabulous solution to the independent researcher's problem of #access to #information.”@deepthiw
“My last article couldn't be possible without the platform @deepdyve that makes journal papers cheaper.”@JoseServera