PART 4: Technology, education and awareness
8: Technology as part of the solution
FREE TRIIODOTHYRONINE ASSAY (FREE T3): The quantitative determination of Free T3
concentration was done by double assay from sera of subjects using a Microplate Enzyme Immunoassay (Batch Number 1325-300 supplied by MonobindInc Ltd, USA). The reference values for normal adults range from 1.4 to 4.2 pg/ml. The sensitivity of this assay was 0.835pg/ml.
The intra assay coefficient of variation (CV) was 3.6%, while the inter assay coefficient of variation (CV) was 7.9% (Appendix V)
FREE THYROXINE ASSAY (FREE T4): The quantitative determination of Free T4
concentration was done by double assay from sera of subjects using a Microplate Enzyme Immunoassay (Batch Number 1225-300 supplied by MonobindInc, USA). The reference value for normal adults ranges from 0.8 to 2.0ng/dl. The sensitivity of this assay was 0.314ng/dl. The intra assay coefficient of variation was 4.26% while the inter assay coefficient of variation was 6.01%
(Appendix IV).
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THYROID STIMULATING HORMONE (TSH) ASSAY: The Quantitative determination of TSH was measured by double assay from sera of subjects using Enzyme Immunoassay Test kits (Batch Number: 10304 supplied by Perfemed Group, Inc Ltd, USA). The reference values for normal adults range from 0.4 to 7.0µIU/ml (Appendix VI).
ANTI-THYROID PEROXIDASE ASSAY (Anti-TPO): Quantitative determination of anti-TPO was measured by double assay from sera of subjects using Enzyme Immunoassay test kits batch number: 521202 supplied by Genway Biotech, Inc, Ltd, USA. The expected reference value for normal adult is less than 30AU/ml. The analytical sensitivity was less than 0.5AU/ml. The intraassay coefficient of variation was 4.8% while the interassay coefficient of variation was 5.7%
(Appendix VII).
ANTI-THYROGLOBULIN ASSAY (Anti-TG): Quantitative determination of anti-TG was measured by double assay from sera of subjects using Enzyme Immunoassay test kits batch number: 521201 supplied by Genway Biotech Inc, Ltd, USA. The expected reference value for normal adults is less than 8AU/ml. The analytical sensitivity is less than 0.5AU/ml. The intraassay coefficient of variation was 1.65% while the interassay precision was 8.5% (Appendix VIII).
FASTING LIPID PROFILE: Total Cholesterol, Low Density Lipoprotein Cholesterol, High Density Lipoprotein-Cholesterol and Triglyceride were measured from the plasma samples by spectroscopy technique using URIT 8020 chemistry Auto-Analyzer (URIT Medical Electronic Co, Ltd, China) with RANDOX Lipid test kits from RANDOX Laboratories Ltd, UK (Appendix VIII).
LABORATORY ANALYSIS OF THYROID HORMONES, TSH AND THYROID AUTOANTIBODIES I
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Figure 3.1: Workbench showing ELISA microplate well, standards and enzyme conjugates for the analysis of thyroid hormones and autoantibodies
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LABORATORY ANALYSIS OF THYROID HORMONES, TSH AND THYROID AUTOANTIBODIES II
Figure 3.2: Workbench showing sera of subjects to be analyzed.
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LABORATORY ANALYSIS OF THYROID HORMONES, TSH AND THYROID AUTOANTIBODIES III
Figure 3.3: Researcher working under the guidance of chemical pathologist.
64 3.15 THYROID ULTRASONOGRAPHY
All subjects had thyroid ultrasonography. Imaging was done by the researcher under guidance of an experienced Radiologist. This was done by placing each subject in a supine position, exposing the neck area of the subject by removing clothing and jewelries around it. The neck of the subject was kept in a slightly extended position allowing the head move above a pillow to expose the neck.
The machine stayed to the right hand side of the patient lying down and to the left hand side of the sonographer. Gel was applied over the side of the neck and a large transducer of 7-15Hz held in transverse and horizontal planes was used to take thyroid images. Each lobe of the thyroid was scanned with visualization of the common carotid and internal jugular vein laterally and the trachea medially. Measurements were taken in the longitudinal and transverse planes using calipers on the ultrasonography machine. The thyroid substance was thoroughly inspected for the presence of cysts, nodules, solid masses and mixed echogenic substances. The thyroid volume was calculated from the measurements obtained using the formula:
The Volume (ml) = 0.479 x d x w x l where d – depth, w – width, l – length(81)
Thyroid ultrasound scan done by the researcher and actively supervised and guided by the Radiologist.
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Figure 3.4: Thyroid ultrasound images done by the researcher under the guidance of a Radiologist
66 Thyroid ultrasound image I
67 Figure 3.6: Thyroid image of a control normal subject
Thyroid ultrasound image II
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Thyroid ultrasound image III
69 3.16 DATA MANAGEMENT AND ANALYSIS
The data obtained were coded and entered into a computer for analysis. Data were analyzed using the Statistical Package for Social Sciences Software (SPSS) version 23 (SPSS, Chicago, IL, WA).
Except where otherwise stated, continuous variables were expressed as mean ± standard deviation (SD) and number count (N) with proportions (%) for categorical data. Descriptive analyses were presented with frequency tables and charts as appropriate.
The statistical analysis method used to compare continuous variables among the three groups was the Analysis of Variance (ANOVA). Thereafter, post-hoc test were carried out on the result of ANOVA tables generated to identify the two groups with a statistically significant difference.
Student’s t-test was used to compare continuous variables when two groups were involved.
70 3.17 DEFINITION OF TERMS
HIV POSITIVE: Subject who have been exposed to the Human Immunodeficiency Virus (HIV) and two HIV tests – preliminary enzyme immunoassay (EIA) test and a confirmatory Western blot are both positive for antibodies to HIV.
HIV NEGATIVE: Subjects who did not have any HIV antibodies discovered in their blood samples using the immunoassay at the point of taking the blood samples with the exception of the
“window period” when the body has not developed any antibodies that can be detected.
ACQUIRED IMMUNODEFICIENCY SYNDROME (AIDS): HIV infection with CD4 count
<200 cells/mm3 and development of specified opportunistic infections and tumorigenic diseases.
OVERT HYPERTHYROIDISM: Defined as elevated serum free Triiodothyronine (FT3) and free thyroxine (FT4) levels in the presence of a very low to undetectable serum TSH levels.
SUBCLINICAL HYPERTHYROIDISM: Defined as a normal FT3 and FT4 in the presence of a low TSH value.
OVERT HYPOTHYROIDISM: Defined as elevated serum TSH value in the presence of low levels of FT4 and FT3.
SUBCLINICAL HYPOTHYROIDISM: Defined as normal FT3 and FT4 levels in the presence of an elevated serum TSH.
ISOLATED LOW FT4: Defined as low FT4 level in the presence of normal serum levels of TSH and FT3.
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ISOLATED LOW FT3: Defined as low FT3 levels in the presence of normal serum levels of TSH and FT4.
CLUSTER OF DIFFERENTIATION (CD4): These are surface markers on T cells which provide targets for immunophenotyping of HIV cells.
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CHAPTER FOUR 4.0 RESULTS