['The Bio-Rad QuantaPhase II radioassay (BR), used for 25 years to measure total folate (TFOL) concentrations for the National Health and Nutrition Examination Survey (NHANES), will be discontinued in 2007. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) or a microbiologic assay (MA) will be used in the future.', 'We measured folate species by LC-MS/MS and TFOL by MA and BR in 327 serum samples.', 'LC-MS/MS measured 5-methyltetrahydrofolic acid (5CH(3)THF; 82%), folic acid (FA; 8%), 5-formyltetrahydrofolic acid (5CHOTHF; 6%), tetrahydrofolic acid (THF; 4%), and 5,10-methenyltetrahydrofolic acid (5,10CH=THF; 0%). The sum of the folate species correlated well with TFOL measured by MA (R(2) = 0.97) and BR (R(2) = 0.91). Compared with LC-MS/MS results, MA and BR values were significantly lower (-6% and -29%, respectively); however, these differences were concentration dependent. The MA almost completely recovered folates added to serum samples except for FA [69% (3%)] and THF [36% (10%)]. The BR underrecovered 5CH(3)THF [61% (9%)] and 5CHOTHF [38% (14%)] and overrecovered 5,10CH=THF [234% (32%)]. Multiple linear regression models with log-transformed data yielded a good fit for converting BR data to MA or LC-MS/MS data and MA data to LC-MS/MS data.'] Do comparison of serum folate species analyzed by LC-MS/MS with total folate measured by microbiologic assay and Bio-Rad radioassay?