title: Quantitative reverse transcription-PCR measurement of thyroglobulin mRNA in peripheral blood of healthy subjects.
abstract: BACKGROUND
  Thyroglobulin mRNA can be detected qualitatively in the peripheral blood of patients with metastatic thyroid cancer, thyroid cancer patients with residual thyroid bed uptake, and individuals with no known thyroid disease with intact thyroid glands by use of a lengthy, highly sensitive extraction technique. To improve and broaden the clinical usefulness of this assay, we developed a quantitative reverse transcription (RT)-PCR assay for thyroglobulin mRNA, using RNA recovered from whole blood with a simplified extraction technique.
  
  
  METHODS
  Whole blood was drawn from 32 healthy subjects in standard EDTA blood collection tubes. Total RNA was extracted from whole blood, using the PUREscript RNA Isolation Kit. RT-PCR using intron-spanning primers was used to quantitatively amplify thyroglobulin mRNA, using the ABI PRISM 7700 Sequence Detection System with a fluorescent-labeled, thyroglobulin-specific oligonucleotide probe. Thyroid RNA calibration curves were created using total RNA recovered from a single nondiseased thyroid gland.
  
  
  RESULTS
  Qualitative RT-PCR demonstrated the presence of thyroglobulin mRNA in the whole blood sample of each healthy subject. The mean concentration of thyroglobulin mRNA detected in these subjects was 433 +/- 69 ng of total thyroid RNA per liter of whole blood (range, 26-1502 ng/L). Overall assay imprecision (CV) was 24% for five samples analyzed 10 times each in separate analytical runs on different days.
  
  
  CONCLUSIONS
  Thyroglobulin mRNA can be accurately detected and quantified in peripheral blood from healthy subjects. This new quantitative technique may improve the clinical utility of circulating thyroglobulin mRNA detection in patients with thyroid disease.
authors.affiliation: Department of Medicine, William Beaumont Army Medical Center, El Paso, TX 79920, USA.
authors.forename: S T
authors.initials: ST
authors.lastname: Wingo
authors.forename: M D
authors.initials: MD
authors.lastname: Ringel
authors.forename: J S
authors.initials: JS
authors.lastname: Anderson
authors.forename: A D
authors.initials: AD
authors.lastname: Patel
authors.forename: Y D
authors.initials: YD
authors.lastname: Lukes
authors.forename: Y Y
authors.initials: YY
authors.lastname: Djuh
authors.forename: B
authors.initials: B
authors.lastname: Solomon
authors.forename: D
authors.initials: D
authors.lastname: Nicholson
authors.forename: P L
authors.initials: PL
authors.lastname: Balducci-Silano
authors.forename: M A
authors.initials: MA
authors.lastname: Levine
authors.forename: G L
authors.initials: GL
authors.lastname: Francis
authors.forename: R M
authors.initials: RM
authors.lastname: Tuttle
mesh_terms: D006801:Humans; D012333:RNA, Messenger / Q000097:blood*; D012016:Reference Values; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012680:Sensitivity and Specificity; D013954:Thyroglobulin / Q000097:blood* / Q000235:genetics
chemical_list: D012333:RNA, Messenger; D013954:Thyroglobulin