Sample preparation module for bacterial lysis and isolation of DNA from human urine.
Silica impregnated polymer monolithic columns may provide a simple method for lysing and extracting DNA from bacteria inside of microfluidic chips. Here we use Escherichia coli as a test organism for a point of care thermoplastic microfluidic module designed to take in a urine sample, mix it with lysis buffer, and perform a hybrid chemical/mechanical lysis and solid phase extraction of nucleic acids from the sample. To demonstrate proof-of-concept, we doped human hematuric urine samples with E. coli at concentrations ranging from 10(1)-10(5) colony-forming units/mL (CFU/mL) to simulate patient samples. We then performed on-chip lysis and DNA extraction. The bacterial DNA was amplified using real-time PCR demonstrating lysis and isolation down to 10(1) CFU/mL. Results were comparable to a commercial kit at higher concentrations and performed better at recovering DNA at lower concentrations.
Kulinski
M Dominika
MD
Department of Mechanical Engineering, Boston University, 44 Cummington St., Boston, MA, USA.
Mahalanabis
Madhumita
M
Department of Biomedical Engineering, Boston University, 44 Cummington St., Boston, MA, USA.
Gillers
Sara
S
Boston University Medical Center, 720 Harrison Ave., Boston 02118 MA, USA.
Zhang
Jane Y
JY
Department of Biomedical Engineering, Boston University, 44 Cummington St., Boston, MA, USA.
Singh
Satish
S
Boston University Medical Center, 720 Harrison Ave., Boston 02118 MA, USA.
Klapperich
Catherine M
CM
Department of Mechanical Engineering, Boston University, 44 Cummington St., Boston, MA, USA.
D001433:Bacteriolysis; D001681:Biological Assay; D003198:Computer Simulation; D004269:DNA, Bacterial / Q000302:isolation & purification*; D004209:Disposable Equipment; D004926:Escherichia coli / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000235:genetics; D006801:Humans; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation* / Q000379:methods*; D008855:Microscopy, Electron, Scanning; D008904:Miniaturization; D010957:Plasmids; D016133:Polymerase Chain Reaction; D011933:Reagent Kits, Diagnostic; D012822:Silicon Dioxide / Q000737:chemistry; D014556:Urine / Q000382:microbiology*
D004269:DNA, Bacterial; D011933:Reagent Kits, Diagnostic; D049452:Green Fluorescent Proteins; D012822:Silicon Dioxide