•Controllable surface groups’ density of MNCs was tuned by adjusting the ratio of APTS-to-MNCs.•Excess APTS input exhibited negative influence on antibody conjugation and MICTS detection.•Steric hindrance of highly branch conformation between surface groups from excess APTS input was found.•The controlled surface functionalization of MNCs is the key for highly efficient and sensitive ICTS.By using well-designed magnetic nanocomposites (MNCs) as probes, immunochromatographic test strip (ICTS) has shown some advantages in sensitivity and quantification of in vitro detection methods. In this study, MNCs were modified with controllable surface carboxyl groups through a facile two-step approach. 3-aminopropyltrimethoxysilane (APTS) was firstly grafted onto MNCs by a sol-gel reaction, followed by amide reaction between amino group of APTS and succinic anhydride. Density of controllable groups and conformation on the surface of MNCs were tuned by adjusting the APTS-to-MNCs ratio. Vibrio parahaemolyticus (VP) was used to examine the effect of surface modification of MNCs on the antibody conjugation and detection performance in ICTS. We also proposed the underlying mechanism based on the AFM analyses of antibody conjugation on two typical functionalized MNCs. It was found that with the increasing of APTS input, lower APTS-to-MNCs ratios (0.03 and 0.3 μL/mg) can improve surface carboxylic groups’ density of MNCs. The optimal APTS-to-MNCs ratio (3 μL/mg) promoted 1.4-fold antibody conjugating efficiency, and 3.4-fold signal-to-noise value in the ICTS for 2 × 105 CFU/mL VP as compared with the ratio of 0.03 μL/mg. Whereas, due to the steric hindrance, the antibody conjugating efficiency and signal-to-noise value at high APTS-to-MNCs ratio (12 μL/mg) respectively decreased 13.9% and 74.7% than the optimal ratio. Accordingly, the controlled surface functionalization of MNCs is the key for highly efficient and sensitive ICTS.To investigate the critical effect of the surface functionalization of the magnetic nanoprobes on the detection performance of immunochromatographic test strip (ICTS), serial functionalized MNCs with controlled carboxyl group density were prepared by adjusting APTS-to-MNCs ratios. Due to the steric hindrance, the antibody conjugating efficiency and signal-to-noise value of ICTS for bacterium at high APTS-to-MNCs ratio (12 μL/mg) respectively decreased 13.9% and 74.7% than the optimal ratio (3 μL/mg).

•A highly sensitive and flexible magnetic nanoprobe labeled immunochromatographic assay (MNP/ICTS) platform for pathogen.•A simple (naked-eye observation) and rapid (~ 10 min) system.•Be able to detect 1.58 × 102 CFU/g Vibrio parahaemolyticus in seafood by flexibly combining with pre-incubation.•The total sample pre-treatment plus MNP/ICTS assay only needs about 4.5 h.A magnetic nanoprobe labeled immunochromatographic test strip (MNP/ICTS) was developed to detect food-borne pathogen Vibrio parahaemolyticus. Specific antibody against V. parahaemolyticus was used as test line by coating onto the nitrocellulose membrane. Magnetic nanoprobe was prepared by immobilizing the specific antibody onto the surface of superparamagnetic nanoparticles. Specificity and sensitivity of the MNP/ICTS system were verified by artificially contaminated shrimp homogenate samples. Reliability and application feasibility of the MNP/ICTS system were demonstrated by using seafood samples (n = 36). Comparing with polymerase chain reaction (PCR) and traditional culture methods, the MNP/ICTS system is found to be not only a rapid qualitative analysis (~ 10 min), but also an accurately quantitative detection platform. Through its rapid magnetic separation property, the MNP/ICTS system is capable to flexibly combine with a sample enrichment and pre-incubation process. This combination makes the qualitative sensitivity for the food samples surged more than 100-fold. A naked-eye observation of 1.58 × 102 CFU/g V. parahaemolyticus was realized. This sensitivity could meet the V. parahaemolyticus test threshold value in many countries. Also, the total sample pre-treatment plus MNP/ICTS assay only needs about 4.5 h. Namely, we can get test results in a day. Hence, the developed MNP/ICTS assay platform is simple, rapid and highly sensitive. It is a flexible test platform for pathogen detection. The favorable comparison with PCR and culture methods further proves that the developed MNP/ICTS is applicable into food-borne pathogen or other areas where a simple, rapid, sensitive and point-of-care analysis is desirable.Scheme of a sample pre-incubation combined with MNP/ICTS assay platform for bacteria V. parahaemolyticus. Seafood homogenate was firstly pre-incubated in SPB media for 3–4 h. Subsequently the enriched samples were mixed well with magnetic nanoprobe, and applied to ICTS assay. V. parahaemolyticus could be qualitatively judged by a naked-eye observation and quantitatively detected by a Magnetic Assay Reader.Download full-size image