Comparative performance of ELISA and dot blot assay for TSH‐receptor antibody detection in Graves’ disease

Zulkarnain, Zulkarnain; Ulhaq, Zulvikar Syambani; Sujuti, Hidayat; Soeatmadji, Djoko Wahono; Zufry, Hendra; Wuragil, Dyah Kinasih; Marhendra, Agung Pramana Warih; Riawan, Wibi; Kurniawati, Siti; Oktanella, Yudit; Aulanni’am, Aulanni’am

doi:10.1002/jcla.24288

Cited by 11 publications

(2 citation statements)

References 12 publications

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“…In a recent study comparing two third-generation immunoassays for the measurement of TSHrAb, a high agreement between the two methods was obtained (kappa; 0.82) [ 22 ]. Another recent study demonstrated slightly better diagnostic performance of third-generation enzyme-linked immunosorbent assay (ELISA) methods than second-generation ones [ 23 ]. Recent studies have also demonstrated the utility of a few newer methods for the measurement of TSHrAb such as one-step third-generation radioimmunoassay and dot-blot method are useful, albeit with modestly lower diagnostic accuracy of the latter method [ 24 , 25 ].…”

Section: Discussionmentioning

confidence: 99%

High Diagnostic Accuracy of Thyroid-Stimulating Hormone (TSH) Receptor Antibodies in Distinguishing Graves’ Disease and Subacute Thyrotoxicosis in the Indian Population

Naga Nitin,

Lakkundi,

S L

et al. 2024

Cureus

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Background and objective: Thyrotoxicosis is a common clinical condition encountered in endocrine practice. Graves’ disease and subacute thyroiditis are the two common causes of thyrotoxicosis and often have overlapping clinical and biochemical features. 99mTc thyroid scintigraphy is the most commonly used confirmatory test to differentiate the two conditions but is not available in the majority of the second-tier cities of India. However, obtaining thyroid stimulating hormone (TSH) receptor antibodies (TSHrAb), another accurate test to differentiate the two conditions, in second-tier cities by outsourcing to labs in major cities is a feasible option nowadays. However, the data on the performance of TSHrAb to differentiate the two conditions in Indian patients is limited. Hence, we have evaluated the diagnostic accuracy of TSHrAb in the Indian population to differentiate Graves’ disease and subacute thyroiditis. Materials and methods: This prospective study was conducted on 115 consecutive newly diagnosed thyrotoxicosis patients presenting to the Department of Endocrinology at a tertiary health care centre in India. Clinical parameters like throat pain, duration of symptoms, and grade of goitre were noted. Measurement of total tri-iodothyronine (TT3), total thyroxine (TT4), TSH, TSHrAb, and 99mTc thyroid scintigraphy were performed in all participants. All participants were followed up at least for six months after the recruitment. Increased tracer uptake (>4%) and/or increased thyroid to parotid trace uptake ratio (>2.5) were used to diagnose Graves’ disease. Results: Eighty-one and 34 patients were diagnosed with Graves’ disease and subacute thyroiditis, respectively. TT3/TT4 ratio had low diagnostic accuracy (area under the curve (AUC): 0.6, best cut-off: 15.6, sensitivity: 53.1%, specificity: 79.4%). TSHrAb had the best AUC (0.9) to distinguish Graves’ disease from subacute thyroiditis (cut-off: 2.0 IU/L, sensitivity: 97.5%, specificity: 100%). In contrast, the kit manufacturer’s reference range (1.75 IU/L) was slightly more sensitive (98.8%), but less specific (94%). Conclusion: The TT3/TT4 ratio is not a good test to differentiate Graves’ disease and subacute thyroiditis. TSHrAb is accurate in distinguishing Graves’ disease from subacute thyroiditis and a level of 2.0 may be a more accurate cut-off to differentiate the two conditions in the Indian population.

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Section: Discussionmentioning

confidence: 99%

High Diagnostic Accuracy of Thyroid-Stimulating Hormone (TSH) Receptor Antibodies in Distinguishing Graves’ Disease and Subacute Thyrotoxicosis in the Indian Population

Naga Nitin,

Lakkundi,

S L

et al. 2024

Cureus

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“…Compared with immunoblotting, ELISA typically has higher sensitivity and throughput, and requires a relatively low amount of materials [24][25][26]. Therefore, we chose ELISA as the detection platform.…”

Section: Optimization Of Elisa Conditions For the Detection Of Tfam-dpcsmentioning

confidence: 99%

An Enzyme-Linked Immunosorbent Assay for the Detection of Mitochondrial DNA–Protein Cross-Links from Mammalian Cells

Zhao

2022

DNA

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DNA–Protein cross-links (DPCs) are cytotoxic DNA lesions with a protein covalently bound to the DNA. Although much has been learned about the formation, repair, and biological consequences of DPCs in the nucleus, little is known regarding mitochondrial DPCs. This is due in part to the lack of robust and specific methods to measure mitochondrial DPCs. Herein, we reported an enzyme-linked immunosorbent assay (ELISA)-based method for detecting mitochondrial DPCs formed between DNA and mitochondrial transcription factor A (TFAM) in cultured human cells. To optimize the purification and detection workflow, we prepared model TFAM-DPCs via Schiff base chemistry using recombinant human TFAM and a DNA substrate containing an abasic (AP) lesion. We optimized the isolation of TFAM-DPCs using commercial silica gel-based columns to achieve a high recovery yield for DPCs. We evaluated the microplate, DNA-coating solution, and HRP substrate for specific and sensitive detection of TFAM-DPCs. Additionally, we optimized the mtDNA isolation procedure to eliminate almost all nuclear DNA contaminants. For proof of concept, we detected the different levels of TFAM-DPCs in mtDNA from HEK293 cells under different biological conditions. The method is based on commercially available materials and can be amended to detect other types of DPCs in mitochondria.

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Comparative performance of ELISA and dot blot assay for TSH‐receptor antibody detection in Graves’ disease

Zulkarnain

Ulhaq

Sujuti

et al. 2022

Clinical Laboratory Analysis

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Background Graves’ disease (GD) is an autoimmune disease, and it accounts for major cases of hyperthyroidism. Antibody against thyroid‐stimulating hormone receptor/TSHR (TRAb) is responsible for hyperthyroidism and is considered as a diagnostic marker for GD. Therefore, we developed a recombinant protein of human TSHR‐169 (hTSHR‐169), which was specifically recognized TRAb in the serum of GD patients and then compare the diagnostic performance between ELISA and dot blot of TRAb tests for their ability to diagnose GD. Methods 20 GD patients and 20 healthy individuals from the Indonesian population were enrolled. TRAb concentration and density were quantified. Comparative analysis was performed using receiver‐operating curve (ROC) analysis. Results For dot blot assay, the minimum concentration to detect TRAb requiring 100 ng of antigen with antiserum diluted at 1:60. For diagnosing GD, the ELISA yielded a higher AUC compared with the dot blot assay (0.95 and 0.85, respectively). Using the recommended cutoff values, the efficiency of both assays was examined by comparing the specificity and sensitivity of the assays to the clinical diagnosis. The ELISA showed 80% and 95%, while the dot blot assay showed 70% and 95% sensitivity and specificity, respectively. Conclusion Although the dot blot assay exhibited lower performance than the ELISA method, the dot blot assay is a simple and rapid diagnostic assay that is suitable for diagnosing GD in rural areas, in which healthcare facilities sometimes are not accessible.

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Comparative performance of ELISA and dot blot assay for TSH‐receptor antibody detection in Graves’ disease

Cited by 11 publications

References 12 publications

High Diagnostic Accuracy of Thyroid-Stimulating Hormone (TSH) Receptor Antibodies in Distinguishing Graves’ Disease and Subacute Thyrotoxicosis in the Indian Population

High Diagnostic Accuracy of Thyroid-Stimulating Hormone (TSH) Receptor Antibodies in Distinguishing Graves’ Disease and Subacute Thyrotoxicosis in the Indian Population

An Enzyme-Linked Immunosorbent Assay for the Detection of Mitochondrial DNA–Protein Cross-Links from Mammalian Cells

Comparative performance of ELISA and dot blot assay for TSH‐receptor antibody detection in Graves’ disease

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