Salivary IGF-1 detection in growing patients with a lateral flow immunochromatographic assay. A proof-of-concept study

This study investigated the potential of a lateral flow immunochromatographic assay (LFIA) for the detection of salivary IGF-1 as a noninvasive biomarker of growth in a sample of 35 orthodontic patients. The patients consisted of 22 females and 13 males, aged between 8 and 16 years. In addition to saliva sampling, patients underwent body mass index (BMI) analysis and a frequently used method for assessing skeletal growth, i.e., the radiographic assessment of skeletal maturation using the third finger Middle Phalanx Maturation method (MPM), which assesses the degree of skeletal growth based on the degree of ossification of the middle phalanx of the third finger on an x-ray. Salivary IGF-1, involved in tissue and skeletal development, was measured using the ELISA method and a prototype of LFIA, the results of which were compared with the MPM method. The following reagents were used to develop the LFIA: anti-IGF1 monoclonal antibody, anti-IGF1 polyclonal antibody, human IGF-1 standard, and protein A. The monoclonal antibody was used to develop the Test line, protein A to develop the Control line, and the polyclonal antibody bound with colloidal gold particles to develop the Conjugate Pad. Sample collection was carried out at the Orthognathodontics clinic at Careggi, Florence, while ELISA analyses and the development of the LFIA prototype were performed at the Department of Clinical and Experimental Medicine at Careggi, Florence. MPM analysis revealed that 9 of 35 patients were at the peak of growth (MPS3), 9 patients were pre-peak (3 in MPS1 and 6 in MPS2), and 17 were post-peak (5 in MPS4 and 12 in MPS5). ELISA analysis revealed mean salivary IGF-1 levels of 3.79 ± 1.94 ng/mL, with higher mean levels observed in MPS3 (4.39 ± 2.01 ng/mL), i.e., at the peak of growth, and lower levels in MPS1 (3.44 ± 1.39 ng/mL), pre-peak of growth, and MPS5 (3.25 ± 1.62 ng/mL), post-peak of growth. The BMI test showed four patients with excess weight problems. 5 Among these, a patient classified as MPS2, whose class had a mean IGF1 value of 3.95 ± 1.77 ng/mL, showed a higher value, 6.44 ng/mL, while another in MPS5 phase (with a mean value of 3.25 ± 1.55 ng/mL), showed a lower IGF1 value, 1.76 ng/mL. The LFIA demonstrated a visual limit of detection between 40 and 80 ng/mL, but positively detected the presence of salivary IGF1 in some MPS3 samples, even with analyte concentrations detected by ELISA above 3.12 ng/mL. Issues related to the matrix effect of saliva and the sensitivity of the test were identified, which indicate the need for further optimization. Although with pending improvements in sensitivity and reproducibility, our preliminary results are promising for assessing the LFIA as a useful tool for a rapid chairside test to optimize the timing of orthodontic procedures.