A simple blood preparation method for nucleic acid amplification tests using membranes.

ABSTRACT

BACKGROUND: To detect most of bloodborne pathogens, serum must be separated from whole blood for efficient nucleic acid amplification. Centrifugation is the most commonly used preparation step for whole blood, but it is not easy to use a centrifuge in rural areas where electricity is not accessible. OBJECTIVE: This study aimed to develop a simple method for obtaining serum suitable for nucleic acid amplification without the use of any instruments. METHODS: Whole blood spiked with Escherichia coli (E. coli) was separated into serum and cellular fraction using 2 closely attached membranes with different characteristics. After brief heating, bacterial DNA in the serum was used for polymerase chain reaction (PCR). RESULTS: Serum was successfully separated from cellular fraction after filtration of one membrane sheet. Membrane sheet containing serum was heated and bacterial DNA in the serum was used for PCR. The quality and concentration of DNA in the heated serum was sufficient for PCR and amplified E. coli gene products were observed. CONCLUSIONS: Separation of bacteria-containing serum was feasible using two membrane sheets and the DNA isolated from serum can be used for PCR after brief heating.