Sensitive and label-free SPR biosensing platforms for high-throughput screening of plasma membrane receptors interactions with insulin-like targets of hypoglycaemic activity

• Published in: Talanta (Oxford) Vol. 274; p. 125914
• Main Authors: Drozd, Marcin, Kobylska, Ewa, Żmieńko, Małgorzata, Chudy, Michał
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01.07.2024
• Subjects: Biosensing Techniques - methods; Equilibrium and kinetic analysis; High-Throughput Screening Assays - methods; Humans; Hypoglycemic Agents - chemistry; Insulin - analysis; Insulin - metabolism; Insulin Glargine - chemistry; Insulin receptor; Protein immobilization; Protein receptor layer; Receptor, IGF Type 1 - metabolism; Receptor, Insulin - metabolism; SPR biosensor; Surface plasmon resonance; Surface Plasmon Resonance - methods; Protein immobilization; Surface plasmon resonance; Insulin receptor; Protein receptor layer; SPR biosensor; Equilibrium and kinetic analysis
• ISSN: 0039-9140; 1873-3573
• DOI: 10.1016/j.talanta.2024.125914

Progress in medical sciences aims for tailored therapy of civilization diseases like diabetes. Preclinical screening of new medicines superior to insulin should include the verification of their affinity to the membrane receptors naturally stimulated by this hormone: insulin receptor isoforms A and B and insulin-like growth factor receptor. Considering that the affinity constants obtained using different experimental conditions are incomparable, it is essential to develop a robust and reliable method to analyze these interactions. The versatile SPR platform developed in this study enables the evaluation of the bioactivity of hypoglycaemic molecules. Thanks to the comprehensive characterization of miscellaneous aspects of the analytical platform, including the design of the SPR biosensor receptor layer, ensuring interaction specificity, as well as the quality control of the standards used (human insulin, HI; long-acting insulin analog: glargine, Gla), the feasibility of the method of equilibrium and kinetic constants determination for insulin-like targets was confirmed. SPR assays constructed in the direct format using IR-A, IR-B, and IGF1-R receptor proteins show high sensitivities and low detection limits towards insulin and glargine detection in the range of 18.3–53.3 nM with no signs of mass transport limitations. The improved analytical performance and stability of SPR biosensors favor the acquisition of good-quality kinetic data, while preservation of receptors activity after binding to long-chain carboxymethyldextran, combined with spontaneous regeneration, results in stability and long shelf life of the biosensor, which makes it useful for label-free insulin analogs biosensing and thus extensive screening in diabetic drugs discovery. [Display omitted] •A label-free SPR platform mimicking living cell biointerface was developed.•Interactions of cell membrane receptors with insulin-like targets can be monitored.•Biosensor shows extended lifespan due to layer stability and regeneration facility.•High sensitivity of label-free SPR biosensors provides reliable determination of KD.•SPR platform enables screening of therapeutic/mutagenic potential of insulin analogs.