Researchers from IIT Guwahati have developed paper-based miniaturized immunosensor for naked eye ALP detection for the first time and have demonstrated its ability for onsite analysis.
Alkaline Phosphatase (ALP) is a metalloprotein found naturally in raw milk samples and is considered as an important biomarker in quality control of milk. Its easy, personalized, as well as instrument-less detection is important to ensure the pasteurization and its differentiation from raw milk.
Doctorate student Kuldeep Mahto under the able guidance of Dr. Pranjal Chandra, Assistant Professor and Ramanujan Fellow have developed an office punching machine crafted paper biosensor for naked eye detection of ALP in milk samples.
The novelty of the work is its design where no instrument was required to design or operate the biosensor and as it was fabricated on just normal cellulose paper, the method is very cheap and commercially viable.
The fabrication of the biosensing probe was characterized using DIC, Fourier transform infrared spectroscopy (FTIR), and atomic force microscopy (AFM). The detection was based on immunocomplexation between the sensor-probe and ALP, which generates blue-green precipitate as an analytical signal by exploiting the catalytic activity of ALP towards 5-bromo-4-chloro 3-indolyl phosphate (BCIP). The dose dependent appearance of the blue-green complex was captured using smartphone camera and DIC was employed using Red, Green, and Blue (RGB) profiling system, where the maximum sensitivity was obtained for the red color channel. Based on the DIC analysis, a wide dynamic range for the ALP detection is obtained from 10-1000 U/mL with the detection limit of 0.87 (±0.07) U/mL. Interferences due to components present in the milk samples were evaluated and the long-term stability of the designed biosensor was examined.
The designed paper-based biosensor is successfully applied to detect ALP in commercial and raw milk samples. Based on the detection principle, a miniaturized kit [20.0 mm (L) x 20.0 mm (W) x 2.15 mm (H)] was developed and applied for the ALP detection to demonstrate the instrument-free direct in-kitchen applicability.
The research was published in the journal Biosensors and Bioelectronics, United Kingdom (Impact Factor 8.17).