NMR systems and methods for the rapid detection of analytes
This invention features systems and methods for the detection of analytes, and their use in the treatment and diagnosis of disease.
1. A method for amplifying a target fungal nucleic acid in a whole blood sample, the method comprising:
(a) contacting a whole blood sample suspected of containing one or more fungal cells with an erythrocyte lysis agent, thereby lysing red blood cells;
(b) centrifuging the product of step (a) to form a supernatant and a pellet;
(c) discarding some or all of the supernatant of step (b) and resuspending the pellet to form an extract;
(d) combining said extract of step (c) with beads to form a mixture and agitating the mixture to form a lysate, said lysate containing both subject cell nucleic acid and fungal nucleic acid; and
(e) providing the lysate of step (d) in a detection tube and amplifying fungal nucleic acids therein by PCR to form an amplified lysate solution;
wherein ten fungal cells per milliliter of said whole blood sample is sufficient to permit amplification of said target fungal nucleic acid.
2. The method of claim 1 , wherein the amplifying of step (e) comprises amplifying a Candida nucleic acid to be detected in the presence of a forward primer and a reverse primer, each of which is universal to multiple Candida species to form a solution comprising a Candida amplicon.
3. The method of claim 1 , wherein the lysing step (a) is by detergent lysis or hypotonic lysis.
4. The method of claim 1 , wherein said amplifying step (e) comprises asymmetric polymerase chain reaction.
5. The method of claim 1 , wherein said amplified lysate solution of step (e) comprises whole blood proteins and non-target oligonucleotides.
6. The method of claim 1 , further comprising (f) detecting said amplified target nucleic acid.