Label-Free Physical Techniques and Methodologies for Proteins Detection in Microfluidic Biosensor Structures

Abstract

Proteins in biological fluids (blood, urine, cerebrospinal fluid) are important biomarkers of various pathological conditions. Protein biomarkers detection and quantification have been proven to be an indispensable diagnostic tool in clinical practice. There is a growing tendency towards using portable diagnostic biosensor devices for point-of-care (POC) analysis based on microfluidic technology as an alternative to conventional laboratory protein assays. In contrast to universally accepted analytical methods involving protein labeling, label-free approaches often allow the development of biosensors with minimal requirements for sample preparation by omitting expensive labelling reagents. The aim of the present work is to review the variety of physical label-free techniques of protein detection and characterization which are suitable for application in micro-fluidic structures and analyze the technological and material aspects of label-free biosensors that implement these methods. The most widely used optical and impedance spectroscopy techniques: absorption, fluorescence, surface plasmon resonance, Raman scattering, and interferometry, as well as new trends in photonics are reviewed. The challenges of materials selection, surfaces tailoring in microfluidic structures, and enhancement of the sensitivity and miniaturization of biosensor systems are discussed. The review provides an overview for current advances and future trends in microfluidics integrated technologies for label-free protein biomarkers detection and discusses existing challenges and a way towards novel solutions.

Optical Chemical Sensor Based on Fast-Protein Liquid Chromatography for Regular Peritoneal Protein Loss Assessment in End-Stage Renal Disease Patients on Continuous Ambulatory Peritoneal Dialysis

“…The angular The graph reveals a good concordance of the FPLC data from the sensor with the protein concentrations measured by the conventional biochemical method; the Pearson correlation coefficient between the FPLC and laboratory results is R 2 = 0.947. The angular coefficient of the linear regression for the dialysate H = 6.65 is higher than the angular coefficient for the BSA solutions H = 4.73, possibly because the dialysate contains not only albumin but also immunoglobulins and other proteins with higher tryptophan content and stronger UV absorption [47,56].…”

“…Earlier, a similar system for the diagnosis of proteinuria was proposed and tested by the authors on urine samples [46]. The working wavelength of the chromatographic detector was chosen according to the characteristic absorption spectra of plasma proteins (mainly albumin) in the spectral range from 250 nm to 290 nm [47]; the absorption spectrum of the bovine serum albumin (BSA) water solution (concentration 2 g/L, cuvette thickness 10 mm) and the normalized spectrum of the UV LED emission measured with the AvaSpec-2048 spectrophotometer from Avantes B.V. (Apeldoorn, the Netherlands) are presented in Figure 1b. The solution was prepared from 99% pure BSA powder (Lot No.…”

“…The sensor consists of the following basic components (Figure 2): 1 is the LabMate buffer reservoir (Code No. 18-3216-03) from GE Healthcare ® Bio-Sciences AB (Uppsala, Sweden); 2 is the chromatographic column PD-10; 3 is the three-way valve for changing liquid flow direction; 4 is the photometric unit for detecting the optical absorption of the eluate at 285 nm; 5 is the drain vessel; 6 is the USB interface cable; 7 is the laptop with the Windows ® operating system and the sensor-dedicated software installed; 8 is the liquid The working wavelength of the chromatographic detector was chosen according to the characteristic absorption spectra of plasma proteins (mainly albumin) in the spectral range from 250 nm to 290 nm [47]; the absorption spectrum of the bovine serum albumin (BSA) water solution (concentration 2 g/L, cuvette thickness 10 mm) and the normalized spectrum of the UV LED emission measured with the AvaSpec-2048 spectrophotometer from Avantes B.V. (Apeldoorn, the Netherlands) are presented in Figure 1b. The solution was prepared from 99% pure BSA powder (Lot No.…”

“…Optical absorption of almost all proteins in the UV region originates from aromatic amino acid residues: tryptophan, tyrosine, and phenylalanine. As a result, protein absorption spectra are very similar having the same characteristic absorption maximum around a wavelength of 280 nm but a relatively high level of variability in specific absorption coefficients, depending on the tryptophan content [47]. It is reasonable to propose that the sensor can determine the total protein concentration in biological fluids, but for each type of sample, separate calibration should be performed because of the possibly different protein compositions.…”

“…In 30-60 s when the sample is fully absorbed by the gel, an additional 25 mL of buffer solution must be introduced into the buffer reservoir and the recording of a chromatogram must be initiated in the sensor software. Over the next 10-15 min, optical transmission of the eluate T(t) at a wavelength of 285 nm is continuously detected with the photometric unit (Figure 2) with a time interval of 1 s. For subsequent processing of the obtained chromatographic data, optical transmission should be recalculated into absorption in accordance with the Bouguer-Lambert-Beer law, the light intensity at the starting point when pure buffer solution is flowing through the cuvette is used as a reference [47]:…”