1. Interferences in hormones imunoassays Facts and Traps Mariana Purice, Andra Caragheorgheopol, Cristina Perhaita, Ecaterina Dumitriu, Florin Alexiu, Corin Badiu “ C.I.Parhon”, National Institute of Endocrinology Bucharest, Romania Balkan Congress for Laboratory Medicine Bucharest, 21-23 September,2011

2. PREAMBUL Diseases are very often not physically evident in endocrinology. Thus clinicians rely on laboratory results to diagnose and monitor patients. To analyse hormones, an ideal tool should allow that starting from a complex, unpurified biological matrix, to quantify compounds with different structure and physical properties. The evolution of techniques over the past 40 years has been focused specifically on indirect methods (such as immuno assay - IA ) where we measure an effect of the biological markers of interest:  For in vitro techniques the effect is the binding of the molecule to a specific antibody, antigen or receptor.

3. Interferences in Immunoassay: Interferences in Immunoassay: In all indirect methods (RIA, ELISA, LIA, CHEMI, etc. generally in immune assays) there are interferences mainly related to: 1. The presence of auto antibodies in the patients serum (ex: anti T3, antiT4, anti insulin, antiTg,etc) 2. Drug/hormonal therapy 3. The presence of macro or micro forms of the native hormone that does not recognize the specific antibody in the immunoassay system.

4. OUR “CASES” We will present 3 specific situations from our practice, for which clinical and research laboratories obtained falsely elevated or falsely low values for a peculiar hormone.

5. CASE 1: Thyroglobuline is “falsely” low ( < 1 ng/ml) in some patients with thyroid carcinoma.

6. A. Clinician point of view A. Clinician point of view The measurement of serum thyroglobulin is used only for assessing response and monitoring treatment. In patients who have been treated with total thyroidectomy and 131iodine ablation, detectable serum Tg (> 2ng/mL) is highly suggestive of residual or recurrent tumour, but could also indicate persistence of a remanent of normal thyroid tissue. There is evidence that anti thyroglobulin antibodies themselves have some value in monitoring response to treatment. Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Thyroglobulin antibody [TGB04+TGB05] - BSA and Azide free (ab80783)

7. B. Laboratory point of view B. Laboratory point of view Endogenous TgAb may interefere in Tg assays, causing either falsely elevated or falsely lowered results,making interpretation difficult. Discrepancy of Tg results measured by radioimmunoassay and immunometric assay may indicate interference by TgAb. Therefore, it is recommended that TgAb be measured at the same time as Tg.

8. OUR EXPERIENCE Nuclear Medicine Laboratory, Institute of Endocrinology: years : 2010 & 2011 n=2030 patients with differentiated thyroid cancer during 131 I treatment * same methods for Tg and Anti Tg evaluation: IRMA (CT), for more than 5 years Anti Tg antibody > 30 IU/ml Negative Tg ( < 1 ng/ml) Anti Tg antibody > 30 IU/ml Negative Tg ( < 1 ng/ml) 128 patients21 patients 125 I -IRMA Anti Tgl Ab kit * 125 I -IRMA Tgl kit * Positive Anti Tg value > 70 IU/mlNormal Tg value < 50 ng/ml ( subject non thyroid carcinoma) Equivoque Anti Tg value: 30-70 IU/ml Normal Tg value < 10 ng/ml (follow up in thyroid carcinoma) < 2 ng/ml ( “ target” value for successful 131 treatment ) Analytical sensibility :6 IU/ml Analytical sensibility :03.ng/ml

9. OUR EXPERIENCE : Serum TgAb interfere in Tg assay giving in 7% of patients FALS negative results for thyroglobuline

10. Conclusions There is an opportunity for clinical chemists to come and talk to endocrinologists. Similarly, it is important that clinicians communicate the type of results that help us most to solve interferences problems in any technique that we use. Clinician Clinical Chemist