Department of Avionics, Indian Institute of Space Science and Technology, Thiruvanathapuram 695547, India.
Biosensors (Basel). 2023 Feb 27;13(3):323. doi: 10.3390/bios13030323.
It is important to isolate exosomes (<150 nm) from biofluid for diagnosis or prognosis purposes, followed by sensing of exosomal proteins. In the present work, exosomes are isolated from human serum by immobilizing on a Screen-Printed Electrode (SPE) followed by electric field lysis and electrochemical impedance spectroscopy (EIS)-based sensing of relevant exosomal proteins (HSP70 and HER2). Upon immobilization of exosomes on the surface, the role of different electrical signals (sinusoidal and square wave) in the lysis of exosomes was studied by varying the frequency and voltage. HSP70 was used for EIS to determine the optimal voltage and frequency for lysing the exosomes. It was observed that the low frequencies and, specifically, sinusoidal signals are ideal for lysing exosomes as compared to square signals. The relative quantity of HSP70 obtained by lysing with different voltages (sinusoidal waveform) was compared using Western blotting. After electric field lysis of the exosome with an optimized signal, HER2, a breast cancer biomarker, was detected successfully from serum by EIS. In the proposed technique, 3.5 × 10 exosomes/mL were isolated from serum. With the limit of detection of 10 pg, the designed cell showed a linear detection of HER2 from 0.1 ng to 1 µg. It was observed from the results that the electric field lysis of exosomes not only plays a significant role in releasing the cargo protein but also improves the sensing of surface proteins associated with exosomes.
从生物体液中分离外泌体(<150nm)对于诊断或预后目的非常重要,然后再检测外泌体蛋白。在本工作中,通过将外泌体固定在丝网印刷电极(SPE)上来从人血清中分离外泌体,然后进行电场裂解和基于电化学阻抗谱(EIS)的相关外泌体蛋白(HSP70 和 HER2)的传感。在外泌体固定在表面后,通过改变频率和电压来研究不同电信号(正弦波和方波)在裂解外泌体中的作用。使用 HSP70 通过 EIS 来确定裂解外泌体的最佳电压和频率。结果表明,与方波相比,低频特别是正弦信号更适合裂解外泌体。使用不同电压(正弦波)裂解后获得的 HSP70 的相对量通过 Western 印迹进行比较。在用优化的信号对外泌体进行电场裂解后,成功地通过 EIS 从血清中检测到乳腺癌生物标志物 HER2。在所提出的技术中,从血清中分离出 3.5×10 个外泌体/mL。设计的细胞具有 10pg 的检测限,可从 0.1ng 到 1μg 线性检测 HER2。从结果中可以看出,外泌体的电场裂解不仅在外泌体释放货物蛋白方面发挥重要作用,而且还改善了与外泌体相关的表面蛋白的传感。