Spectroscopic properties of various blood antigens/antibodies

Abstract

Since the traditional method generates biological waste, there is a significant demand for an easy, quick technique of blood type identification without contamination. In fact, individuals can be divided into four main blood groups whose antigens are available in red blood cell (RBC) membranes and the antibodies in the plasma. Here, UV-vis and photoluminescence (PL) spectroscopic methods are systematically used to find the spectra of blood typing antigens (A, B and AB) and antibodies i.e. A-Anti, B-Anti, AB-Anti and D reagent. The PL spectra of RBCs in different blood groups as well as the corresponding antibodies are successfully resolved for the purpose of blood typing. The unique photophysical characteristics of these biomolecules including signal intensity and peak emission wavelength in PL spectra are lucidly anticipated to accurately discriminate ABO groups. PL spectra of RBC in positive blood typing indicate larger signal and shorter emission peak wavelength corresponding to negative ones. Furthermore, the monoclonal antibody PL emissions emphasize that Anti-A benefits higher intensity and shorter peak wavelength (blue shift) than B-Anti. In the following, lucid blue shifts are obtained in terms of antibody concentrations accompanying the elevation of fluorescence signal, most likely due to the aggregation induced emission (AIE) phenomenon, quite the opposite of the aggregation-caused quenching (ACQ) that is widely observed from conventional chromophore. Those are envisaged as unique properties of each antibody to utilize in the spectral blood typing.

Fig. 1.

Chemical structure of various blood antigens and antibody. a) O antigen b) A antigen c) B antigen d) Schematic of a typical antibody containing FC and Fab domains including CDRs and FRs in heavy (VH) and light (VL) chains. Note that amino acid ring residues such as Trp, Tyr and Phe are sequentially located in CDRs. Figure 1 right illustrates the components of whole blood after centrifuging.

Fig. 2.

(a) PL spectra of the red blood cell (including hemoglobin and antigen) at λ_ex=410 nm. (b) Spectral absorbance of Porphyrin with an absorbance wavelength peak of λex=410 nm and the corresponding fluorescence spectrum from 500 to 750 nm [31] (c) PL signal and (d) Peak emission wavelength for various red blood cells.

Fig. 3.

(a) UV-Vis absorbance spectra of the various blood typing antibodies, dominant peak takes place at UV region. (b) Absorbance spectra of Trp, Tyr and Phe amino acid residues [29] (c) Absorbance intensity around 260 nm for antibodies (d) Corresponding peak wavelength of antibodies. Note that these data are unique and reproducible as the main features of each antibody.

Fig. 4.

(a) PL spectra of various blood typing antibodies at λ_ex = 260 nm. (b) Fluorescence spectra of Trp, Tyr and Phe amino acids residues [29] (c) PL emission intensity (d) Corresponding peak wavelength of antibodies. Note that the Trp fluorescence emission resembles to be dominant due to presence of fluorescence peaks around 343-345.5 nm.

Fig. 5.

The PL signal intensity in terms of concentrations (1, 2.5, 5, 7.5 and 10 mg/ml). (a) A-Anti (b) B-Anti. Signal amplitudes and spectral shift versus each antibodies concentration are depicted in up inset and down inset graphs respectively.

Fig. 6.

The PL signal intensity in terms of concentrations (1, 2.5, 5, 7.5 and 10 mg/ml). (a) AB-Anti (b) D-Anti reagent. Signal amplitudes and spectral shift versus each antibodies concentration are depicted in up inset and down inset graphs respectively.

Fig. 7.

(a) PL signal and (b) peak wavelength of A-Anti, B-Anti and AB-Anti versus concentrations (1, 2.5, 5, 7.5 and 10 mg/ml) in saline solution.

Fig. 8.

Block diagram of blood typing based on spectrophotometric approach and traditional clinical method.

Fig. 9.

Common clinical procedure a) forward b) reverse blood typig versus PL spectroscopy of antigens and antibodies as alternative methods of blood typing. Note that AB blood type contain no antibodies, therefore no signal is detected.