Modern Approaches to Quality Evaluation of Enzyme-Linked Immunoassay Diagnostic Kits


  • Valeria Shchetinin Weston Foods Inc., Canada



ELISA kits, Diagnostic quality, Sera panels, Control materials


Background. Enzyme-linked immunoassay (ELISA) diagnostic kits are a medical device designed to solve an important task as patient's life and health depend on the correctness of laboratory test result. Therefore, at present, special attention is given to the quality of in vitro diagnostic medical devices, which in turn is related to standardization and technical regulation.

Objective. The goal of the work was analysis of up-to-date methodological approaches to quality assessment of ELISA diagnostic kits.

Methods. The results of the development, standardization, and testing of ELISA kits, as well as the recommendations of international institutions and certification bodies, were studied and analyzed.

Results. Properties of biological components of diagnostic kits (immunoenzymatic conjugates and immunosorbents) were analyzed; their impact on diagnostic quality level of ELISA kits was determined. The current methodology for assessing the quality conjugates and immunoassay diagnostic kits, in general, was analyzed. The important role of standardized sera panels (qualification, verification, seroconversion, expert, and sensitivity panels) in the assessment procedure for the diagnostic ELISA kits has been demonstrated. The methods of obtaining control materials used to assess the quality of diagnostic kits were analyzed. Special attention was paid to methods to ensure the stability of control materials.

Conclusions. In the case of the most socially important infections (HIV, hepatitis B and C), strict values of specificity and sensitivity have been established by regulatory authorities and/or international institutions. In the case of other diseases, rationing of diagnostic indicators has to be performed (justified) by research and development group and/or by the manufacturer.



Galkin OY, Komar AG, Pys'menna MO. Specificity of manifacturing process validation for diagnostic serological devices. Biotechnologia Acta. 2018;11(1):25-38. DOI: 10.15407/biotech11.01.025

Galkin OY, Komar AG, Grygorenko AA. Bioanalytical standardization of medical devices for serological diagnosis. Biotechnologia Acta. 2015;8(2):112-9.

Lutsenko TN, Kovalenko MV, Galkin OY. Validation of biological activity testing procedure of recombinant human interleukin-7. Ukr Biochem J. 2017;89(1):82-9. DOI: 10.15407/ubj89.01.082

Galkin AY, Dugan AM. Elaboration of immunoenzymatic test-kit for total human IgE assay and investigation of its analytical properties. Int J Immunol. 2013;1(1):1-6.

Kabernyuk AA, Oliynyk OS, Redchuk TA, Romanyuk SI, Kolibo DV, Yevtushenko VV, et al. Development of immunochemical test systems for the control of anti-diphtheria immunity in people population. Sci Innov. 2008;4(3):22-31.

Siromolot AA, Redchuk TA, Solodiankin OS, Kolibo DV, Gerilovich AP, Komisarenko SV. The trial of experimental test system for the specific diagnostics of cattle tuberculosis. Biotechnologia Acta. 2016;9(4):14-8. DOI: 10.15407/biotech9.04.014

Galkin OY, Besarab AB, Lutsenko TN. Characteristics of enzyme-linked immunosorbent assay for detection of IgG antibodies specific to Chlamydia trachomatis heat shock protein (HSP-60). Ukr Biochem J. 2017;89(1):22-30. DOI: 10.15407/ubj89.01.022

Linghu X, Segraves NL, Abramovich I, Wong N, Müller B, Neubauer N, et al. Highly efficient synthesis of a Staphylococcus aureus targeting payload to enable the first antibody-antibiotic conjugate. Chemistry – A European Journal. 2018;24(12):2792. DOI: 10.1002/chem.201706013

Golub N, Levtun I. Impact of sound irradiation on Chlorella vulgaris cell metabolism. Eastern-European J Enterprise Technol. 2016;2(10):27-31. DOI: 10.15587/1729-4061.2016.63730

Saegerman C, De Waele L, Gilson D, Godfroid J, Thiange P, Michel P, et al. Evaluation of three serum i-ELISAs using monoclonal antibodies and protein G as peroxidase conjugate for the diagnosis of bovine brucellosis. Vet Microbiol. 2004;100(1-2):91-105. DOI: 10.15587/1729-4061.2016.63730

Hermanson GT. Bioconjugate techniques. 3rd ed. San Diego: Academic Press; 2013.

Oh JG, Seong J, Han S, Heo TH. Development of an anti-EPO antibody detection kit based on lab-on-a-chip and bridging antibody technologies. Biologicals. 2018;54:8-12. DOI: 10.1016/j.biologicals.2018.05.005

Nikolaenko IV, Galkin AY, Raevskaya GE, Kas'ianenko TV, Nereshchenko MI, Donskaia ES, et al. Preparation of monoclonal antibodies to the Fc-fragment of human IgG and the use of their based immunoenzyme conjugates. Clin Lab Diagn. 2005;11:8-11.

Donskaya ES, Raevskaya GE, Nikolaenko IV. The use of conjugate of monoclonal antibodies with polymer peroxidase as a way to increase the sensitivity and reduce the hook effect of high doses in ELISA for the determination of HBsAg. Viral hepatitis B – diagnosis, treatment and prevention. In: Proc. Rus Sci Conf; 2004; Moscow. p. 49-50.

Semynozhenko VP, Moskalenko VF, Hural AL, editors. HIV/AIDS: problems of etiology, epidemiology and diagnosis. Kyiv: Poligraf Express; 2004.

WHO. Recommendations for the preparation, characterization and estab-lishment of international and other biological reference standards. WHO Technical Report Series. 2004.

In vitro diagnostic medical devices – Measurement of quantities in samples of biological origin, description of reference materials. ISO 15194:2009.

Cauchard J, Soldan A, Madeline A, Johnson P, Büscher P, Petry S. Inter-laboratory ring trials to evaluate serological methods for dourine diagnosis. Vet. Parasitol. 2014;205(1-2):70-6. DOI: 10.1016/j.vetpar.2014.06.025

Kubanova AA, Frigo NV, Rotanov SV. System of external and internal quality control of laboratory diagnosis of sexually transmitted diseases in the Russian Federation. Moscow: TsNIKVI; 2006.

National HIV testing day and new testing recommendations. MMWR Morbidity and Mortality Weekly Report. 2014;63(25):537. DOI: 10.15585/mmwr.mm6825a1

Galkin OY. Approaches to the synthesis of conjugates for enzyme immunoassay test-systems and evaluation of their use for diagnostics of infectious diseases. Ukr J Clin Lab Med. 2010;5(4):54-60.

The official website of the Institute Paul Ehrlich behalf of the Federal Ministry of Health of Germany [Internet]. 2019 [cited 2019 Dec 14]. Available from:

Commission decision of 27 November 2009 on common technical specifications for in vitro diagnostic medical devices (2009/886/EC). Official Journal of the European Union. 2009;L318:25-40.

Parreno V, Romera SA, Makek L, Rodriguez D, Malacari D, Maidana S, et al. Validation of an indirect ELISA to detect antibodies against BoHV-1 inbovine and guinea-pig serum samples using ISO/IEC 17025 standards. J Virol Meth. 2010;169(1):143-53. DOI: 10.1016/j.jviromet.2010.07.014

Ederveen JC. A practical approach to biological assay validation. Hoofddorp: Progress; 2010.

Schultheiss OC, Stanton SJ. Assessment of salivary hormones. Methods in Social Neuroscience. New York: Guilford Press; 2009.

Ricos C, Juvany R, Simon M, Hernández A, Alvarez V, Jiménez CV, et al. Commutability and traceability: their repercussions on analytical bias and inaccuracy. Clin Chim Acta. 1999;280(1-2):135-45. DOI: 10.1016/s0009-8981(98)00187-9

Hu M, Liu N, Cai YY, Lei T, Zhang M. Accuracy analysis and comparison of different serological detection methods in syphilis. Zhonghua Yi Xue Za Zhi. 2017;97(36):2844-7.

Ivchenko SN, Susloparov IM, Masyicheva VI. Construction of a liquid panel of sera containing and not containing antibodies of IgG class to human cytomegalovirus. Vestnik RAMN. 2004;8:37-40.

Castro AR, Kikkert SE, Fears MB, Pope V. Defibrination of blood plasma for use in serological tests for syphilis. Clin Diagn Lab Immunol. 2002;9(6):1376-8. DOI: 10.1128/cdli.9.6.1376-1378.2002

Kanev AN, Vorobeva MS, Shalunova NV. Development of standard serum panels for quality control of immunoenzyme test systems in Russia. Vestnik RAMN. 1998;3:47-51.

Jeffcoate SL. WHO guidelines for the preparation of international standards and other reference materials for biological substances. Dev Biol Stand. 1992;74:195-201.

Pankratov O, Shimanskaya I, Pankratov V, Navrotsky A, Ballard RC, Unemo M, et al. Laboratory diagnosis of sexually transmitted infections in Belarus. Acta Derm Venereol. 2011;91(1):64-5. DOI: 10.2340/00015555-0982

Henoh MA, Pershina VP, Lapinskaya EM. Effect of deep freezing on protein solutions. Tsitologiya. 1966;8(6):769-72.

Schellekens H, Casadevall N. Immunogenicity of recombinant human proteins: causes and consequences. J Neurol. 2004;251(2):4-9. DOI: 10.1007/s00415-004-1202-9

MacLennan S, Barbara J. Risks and side effects of therapy with plasma and plasma fractions. Best Pract Res Clin Hematol. 2006;19(1):169-89. DOI: 10.1016/j.beha.2005.01.033

Kostantino GR, Shvenderman SR, Langer R. Damage to preparations of lyophilized protein. Biochemistry (Moscow). 1998;63(3):422-9.

Lock RJ. My approach to internal quality control in a clinical immunology laboratory. J Clin Pathol. 2006;59(7):681-4.

Kanev AN, Vorobeva MS, Shalunova NV. The construction of standard serum panels with a normalized IgG antibody level. Voprosyi Virusologii. 1996;41(4):161-6.

Kanev AN, Shalunova NV, Netesov SV. Designing a reference panel of sera for hepatitis C virus with a normalized level of IgG antibodies. Voprosyi Virusologii. 2000;45(4):42-7.

Lichstein HC, Malcolm HS. Studies of the effect of sodium azide on microbic growth and respiration. J Bacteriol. 1943;47(3):221-30.

Kurashvili LV, Besedina NF. Evaluation of liquid human serum used to control the quality of clinical and biochemical studies. Clin Lab Diagn. 1995;1:6-8.

Costantino H, Schwenderman S, Griebenov K. The secondary structure and aggregation of lyophilized tetanus toxoid. J Pharm Sci. 1996;85(12):1290-3.

Crotts G, Park T. Stability and relese of bovine serum albumin encapsulated with poly (D,L-lactice-co-glycolide) microparticles. J Control Release. 1997;44(1-2):123-34. DOI: 10.1016/s0168-3659(96)01511-8

Sola-Penna M, Meyer-Fernandes J. Stabilisation ageinst thermal inactivation promoted by sugars on enzyme structure and function: why is trehalose more effective then other sugars. Arch Biochem Biophis. 1998;360(1):10-4. DOI: 10.1006/abbi.1998.0906

Imamura K, Ogawa T, Sakiyama T. Effects of types of sugar on the stabilization of protein in the dried state. J Pharm Sci. 2003;92(2):266-74. DOI: 10.1002/jps.10305

Brande J, Landkjager L. Chemical stability of insulin. 3. Influence of excipients, formulations, and pH. Acta Pharm Nord. 1992;4(3):149-58.

Hsieh SY, Chen RK, Pan YH. Systematical evaluation of the effects of sample collection procedures on low-molecular-weight serum/plasma proteome profiling. Proteomics. 2006;6(10):189-98. DOI: 10.1002/pmic.200500535




How to Cite

Shchetinin V. Modern Approaches to Quality Evaluation of Enzyme-Linked Immunoassay Diagnostic Kits. Innov Biosyst Bioeng [Internet]. 2020Jan.30 [cited 2021Sep.22];4(1):4-12. Available from: