Clinical Education Center
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- ABL Kinase Domain Mutation in CML, Cell-based
- ABO Group and Rh Type
- Acid-Fast Bacillus (AFB) Identification, Sequencing and Stain, Paraffin Block
- ADAMTS13 Activity with Reflex to ADAMTS13 Inhibitor
- Alcohol Metabolites, Quantitative, Urine
- Alpha-Globin Common Mutation Analysis
- Alpha-Globin Gene Deletion or Duplication
- Alpha-Globin Gene Sequencing
- Anti-Müllerian Hormone AssessR™
- Anti-PF4 and Serotonin Release Assay (SRA) for Diagnosing Heparin-induced Thrombocytopenia/Thrombosis (HIT/HITT)
- Antiphospholipid Antibodies
- ASCVD Risk Panel with Score
- Autoimmune Epilepsy Evaluation
- Autoimmune Diseases, Tests for
- B-cell and T-cell Clonality Assays by PCR
- B-Type Natriuretic Peptide (BNP)
- BCR-ABL1 Gene Rearrangement, Quantitative PCR
- Beta-Globin Complete
- BRCAvantage®, Ashkenazi Jewish Screen
- BRCAvantage®, Rearrangements
- BRCAvantage™, Comprehensive
- BRCAvantage™, Single Site
- CDH1 Sequencing and Deletion/Duplication
- Clostridium difficile Diagnostic Testing
- C1 Inhibitor, Protein and Functional Tests
- Calreticulin (CALR) Mutation Analysis
- Carbapenem Resistant Enterobacteriaceae Culture Screen
- Cardio IQ Lipoprotein Fractionation, Ion Mobility
- Cervical Cancer, TERC, FISH
- CFvantage® Cystic Fibrosis Expanded Screen
- Chlamydia trachomatis, TMA
- Chlamydia trachomatis/Neisseria gonorrhoeae RNA, TMA
- Chromosomal Microarray, POC, ClariSure®, Oligo-SNP
- Chromosomal Microarray, Postnatal, ClariSure® Oligo-SNP
- Chromosome Analysis and AFP with Reflex to AChE, Fetal Hgb, Amniotic Fluid
- Chromosome Analysis, Amniotic Fluid
- Chromosome Analysis, Blood
- Chromosome Analysis, Blood with Reflex to Postnatal, ClariSure® Oligo-SNP Array
- Chromosome Analysis, Chorionic Villus Sample
- Chromosome Analysis, High Resolution
- Chromosome Analysis, High Resolution with Reflex to Postnatal, ClariSure® Oligo-SNP Array
- Chromosome Analysis, Mosaicism
- Chromosome Analysis, Neonatal Blood
- Chromosome Analysis, Sister Chromatid Exchange
- Chromosome Analysis, Tissue
- Chromosome DEB Assay for Fanconi anemia
- Chronic Lymphocytic Leukemia (CLL) - Diagnostic and Prognostic Testing
- Culture, Fungus
- Culture, Urine, Routine
- Cystic Fibrosis Screen
- Cytomegalovirus (CMV) and Epstein Barr Virus (EBV) PCR
- D-Dimer, Quantitative
- Dementia, Secondary Causes
- Dengue Virus Testing
- Diabetes Risk Panel with Score and Cardio IQ® Diabetes Risk Panel with Score
- Drug Testing, General Toxicology (Blood, Urine, or Serum)
- Drug Toxicology Alcohol Metab, QN, Oral Fluid
- Drug Toxicology Monitoring, Oral Fluid Testing
- Factor V (Leiden) Mutation Analysis
- Familial Mediterranean Fever Mutation Analysis
- First Trimester Screen, hCG
- First Trimester Screen, Hyperglycosylated hCG (h-hCG)
- FISH, Angelman
- FISH, MET Amplification
- FISH, Myeloma, 17p-, rea 14q32 with Reflexes
- FISH, Prader-Willi
- FISH, Prenatal Screen
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- HCV Genotyping
- Helicobacter pylori (H pylori) Antibody Discontinuation
- Heparin, Anti-Xa
- Hepatitis B Surface Antibody, Quantitative
- Hepatitis C Antibody with Reflex to HCV RNA, PCR with Reflex to Genotype
- Hepatitis C Viral RNA Genotype 1 NS5A Drug-resistance
- Hepatitis C Viral RNA Genotype 3 NS5A Drug Resistance
- Hepatitis C Viral RNA NS3 Drug Resistance
- Hepatitis C, RNA, Quantitative, PCR
- Hereditary Cancer Panels: MYvantageTM Hereditary Comprehensive Cancer Panel and GIvantageTM Hereditary Colorectal Cancer Panel
- Hereditary Hemochromatosis DNA Mutation Analysis
- Herpes Simplex Virus (HSV) Type-Specific IgG Antibodies
- Herpes Simplex Virus Type 2 (HSV-2) IgG Inhibition, ELISA
- HIV-1 Coreceptor Tropism, Proviral DNA
- HIV-1 Coreceptor Tropism, Ultradeep Sequencing
- HIV-1 Integrase Genotype
- HIV-1/2 Antigen and Antibodies, Fourth Generation, with Reflexes
- HPV mRNA E6/E7
- Influenza A and B Antigen, Immunoassay
- Influenza Type A and B Antibodies
- Insulin, Intact, LC/MS/MS
- Integrated Screen, Part 1
- Integrated Screen, Part 2
- Intrinsic Factor Blocking Antibody
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- Maternal Serum AFP
- Melanoma, BRAF V600E and V600K Mutation Analysis, THxID®
- Metanephrines, Fractionated, Free, LC/MS/MS, Plasma
- Methylenetetrahydrofolate Reductase (MTHFR), DNA Analysis
- Microalbumin (Urinary Albumin Excretion)
- Pain Management and CYP2D6/CYP2C19
- Pain Management, Naltrexone, Quantitative, Urine
- Partial Thromboplastin Time, Activated (aPTT)
- Penta Screen
- PIK3CA Mutation Analysis
- PNH with FLAER (High Sensitivity)
- Prothrombin Time with INR
- PTH, Intact and Calcium
- Streptococcus pneumoniae (Pneumococcal) Antibody Tests
- Saccharomyces cerevisiae Antibodies (ASCA) (IgG, IgA)
- Sequential Integrated Screen, Part 1
- Sequential Integrated Screen, Part 2
- Serum Integrated Screen, Part 1
- Serum Integrated Screen, Part 2
- Serum Pregnancy Tests
- Sickle Cell Screen
- Stepwise, Part 1
- Stepwise, Part 2
- SureSwab® Trichomonas vaginalis RNA, Qualitative TMA
- SureSwab®, Candidiasis, PCR
- TP53 Sequencing and Deletion/Duplication
- T4, Free
- Tamoxifen and Metabolites, LC-MS/MS
- Testosterone Testing
- Total Testosterone, LC/MS/MS
- Triple Screen
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T4, FreeTest code(s) 35167, 866
Question 1. How many different free T4 tests do you have?
We currently offer two tests for free T4 measurement. One test uses a direct equilibrium dialysis method (T4, Free, Direct Dialysis, test code 35167). The other test measures free T4 in an automated immunoassay on the Siemens Centaur™ platform (T4, Free, test code 866).
Question 2. How is free T4 measured using the direct equilibrium dialysis? What are the performance characteristics?
In the direct equilibrium dialysis (ED) method, patient serum is dialyzed against a buffer for 16-18 hours. ED separates the free T4 molecules from the protein bound T4 molecules. The smaller free T4 molecules can cross the dialysis membrane, but the larger bound T4 molecules cannot. After dialysis, free T4 can be measured using either immunoassay or mass spectrometry. We currently use a sensitive radioimmunoassay that has an analytical sensitivity of 0.2 ng/dL. Results are independent of T4-binding protein concentrations and are unaffected by the presence of molecular variants of these proteins or by circulating thyroid autoantibodies.
Question 3. What can cause an unexpectedly high free T4 result in the dialysis method?
Unexpected high free T4 results by ED may be seen in sera from heparin-treated patients. Free fatty acids (FFA), which can displace T4 from its binding proteins, are generated in these samples. The spuriously high free T4 result appears to be primarily an “in vitro” effect and does not reflect the true free T4 level in vivo.
Less commonly, impurities in the tracer may cause erroneously high results.
If you feel that the results are elevated beyond what was anticipated, please contact the lab to discuss your results.
Question 4. How is free T4 measured using immunoassay?
The immunoassay is performed on the automated Siemens Centaur platform. This assay is a competitive immunoassay using a T4 analog. It assumes that the T4 binding capacity is normal; therefore, results can be impacted when the albumin or thyroid binding globulin (TBG) levels are abnormal. The analytical measurement range is 0.1 – 11 ng/dL. The advantage of this assay is that it provides a shorter turnaround time than does the direct equilibrium dialysis method.
Question 5. What factors can cause inaccurate free T4 results in the immunoassay?
The analog-based immunoassays perform poorly at extremes of thyroid hormone binding capacity. Thus, excess TBG, as seen in select genetic conditions, in pregnancy, and in other conditions with increased estrogen, can cause a slight positive bias for free T4 results. Genetically-related TBG deficiency can also cause a negative bias in the free T4 results for this method. Very low levels of albumin, as seen in critical illness, premature infants, etc., also alter free T4 measurements. Free T4 results tend to be low in patients with abnormally low albumin levels and high in patients with familiar dysalbuminemic hyperthyroxinemia or in those having thyroid hormone autoantibodies.
Heterophilic antibodies in patient serum may also cause erroneous results.
Question 6. How do free T4 results generated from the Centaur immunoassay compare with those generated from direct equilibrium dialysis?
ED is considered by many as the “gold standard” method based on the consistent log-linear relationship seen between free T4 ED measurements and TSH in patients with stable thyroid status.1 Analog-based immunoassays performed on automated platforms provide a convenient estimation of free T4, but results do not always agree with the gold standard ED methods. We found that free T4 results from the Centaur immunoassay were generally 40% to 60% lower than those from ED.2 Linear regression analysis of our data gave a slope of 0.46 and an intercept of 0.05 (R2 =0.79).2 This is consistent with previous observations that immunoassays perform reasonably well for samples with normal free T4 but are less reliable in samples with abnormal free T4.3 We suggest free T4 measurements from the automated immunoassay be repeated using the ED method when they are inconsistent with the clinical picture.
- The National Academy of Clinical Biochemistry. Laboratory Medicine Practice Guidelines. Laboratory Support for the Diagnosis of Thyroid Disease. 2002. http://www.aacc.org/sitecollectiondocuments/nacb/lmpg/thyroid/thyroid-fullversion.pdf. Accessed June 26, 2012.
- Song L, Furlanetto R, Lara M, et al. Comparison of two free T4 immunoassays with a direct dialysis method. Clin Chem. 2011;57(10):A189. http://www.aacc.org/events/annualmtgdirectory/Documents/AACC_11_Abstr_Suppl_A177-A196.pdf. Accessed June 26, 2012.
- Sapin R, d’Herbomez M. Free thyroxine measured by equilibrium dialysis and nine immunoassays in sera with various serum thyroxine-binding capacities. Clin Chem. 2003;49:1531-1535. http://www.clinchem.org/content/49/9/1531.full.pdf+html. Accessed June 26, 2012.