The Detection Of Caspase-3 By Raman Spectroscopy

Apoptosis is one of the mechanisms of cell death and plays an important role in maintaining intracellular and extracellular balance.The abnormalities in cell death regulation during the apoptosis process can cause many severe diseases such as autoimmune diseases,neurodegenerative diseases,and ischemia-related injuries.Therefore,the construction of a rapid and sensitive method to detecting apoptosis is of great significance for scientific research.The process of apoptosis occurs through a complex signaling cascade that is tightly regulated with multiple sites.Caspase proteases family is supposed to play a vital role in the apoptosis and caspase-3 is regarded as one of the most critical proteins.Therefore,monitoring of apoptosis can be achieved by detecting the apoptosis enzyme caspase-3.The study proposed a novel SERS method for sensing caspase-3 with gold nanoparticles(AuNPs)-peptide probe in living cells with high selectivity and sensitivity.Raman reporter(4-MB A)and peptides were anchored on the AuNPs surface through Au-S bond with a high affinity.Peptides CCALNNPFFDEVD and CCALNNKYDDEVD were prepared to attach AuNPs,and the CALNN sequence was used to stable AuNPs and PFF or KYD were acted as assembly parts and DVED was the substrate to be cleaved by caspase-3.In the existence of the caspase-3,the DVED can be cut and PFF or KYD was exposed and the PFF or KYD terminated AuNPs can be assembled to form colloidal system for SERS detection.So,the correlation of intensity of SERS with the amount of caspase-3 can be investigated.The results showed that the linear range of AuNPs-peptide ? was 0-0.2 ?g/mL,the minimum detection limit was 1.5 ng/mL and the correlation coefficient was R2=0.998;the linear range of AuNPs-peptide ? was 0-0.1 ?g/mL,the minimum detection limit was 2.75 ng/mL and the correlation coefficient was R2=0.999.The results showed that the detection limit of AuNPs-peptide ?was lower and the detection range was wider.Finally,this caspase-3 specified AuNPs-peptide? probe showed application in analysis of caspase-3 in live cells.The detection of apoptotic enzyme caspase-3 was realized by Raman spectroscopy.This method has the advantages of high sensitivity,high specificity and simple operation.It provides a theoretical basis and method for the detection of apoptosis and the application of this new probe in the detection of food safety,biomedical and clinical monitoring.