Methods for isolating a microbial analyte

A method for selectively isolating a microbial cell analyte from a sample, the method comprising:

• (a) providing a sample containing mammalian cells and potentially containing microbial cells;
• (b) mixing the sample with a first solution that selectively lyses the mammalian cells to obtain a first lysate containing lysed mammalian cells and intact microbial cells, if present, wherein the first solution is a buffered solution comprising a chaotropic salt that is guanidine hydrochloride, and a detergent that is saponin;
• (c) filtering the first lysate through a filter having a pore size that retains the intact microbial cells, wherein the pore size from about 0.1 μm to about 1 μm;
• (d) contacting the filter containing the retained microbial cells with a second solution that is effective to allow release of the microbial cells from the filter, wherein the second solution is a buffered solution comprising sodium hydroxide at a concentration of from about 10 mM to about 100 mM, dimethyl sulfoxide (DMSO) at a concentration of from about 10% (v/v) to about 20% (v/v), glycerol at a concentration of from about 1% (v/v) to about 15% (v/v), and a chelating agent;
• (e) incubating the filter with the second solution at an incubation temperature of from about 70° C. to about 99° C. to facilitate the release of the microbial cells from the filter;
• (f) collecting the microbial cells from the filter, wherein said collection step comprises
• (i) using an elution buffer to elute the microbial cells from the filter, wherein the direction of fluid flow through the filter is opposite the direction of fluid flow applied in the filtering step (c);
• or
• (ii) aspirating the microbial cells from the filter, wherein the microbial cells are aspirated from the same side of the filter initially contacted by the first lysate at step (c);
• and
• (g) lysing the microbial cells collected at step (f) to obtain a second lysate comprising an analyte released from the microbial cells, thereby selectively isolating a microbial cell analyte from the sample.