Integrated Antibody With Metal-Organic Framework For Immunoassay

In recent years,enzyme-linked immunosorbent assay(ELISA)has been widely used as a gold standard in biomedical field.However,some inevitable shortcomings of still exist in its practical application,especially in the preparation of enzyme-labeled antibody,which involves is a complicated covalent binding.Metal-organic framework(MOF)is a fascinating hybrid material and has been widely examined in a variety filds,such as gas storage,gas adsorption,and drug delivery,due to its large surface area,high porosity and adjustable structure.In this paper,based on the excellent performance of MOF as a carrier,we have developed a series of novel enzyme-labeled antibodies and investigated their applications in immunoassay.The main works as follows:(1)By integrating rabbit anti-mouse immunoglobulin G antibody(RIgG)with copper-based metal-organic framework(Cu-MOF),an enzyme-labeled antibody(RIgG@Cu-MOF)loaded with rabbit anti-mouse immunoglobulin G antibody(RIgG)were prepared.It was found that the specific recognition performance and efficiency of RIgG to its antigen wsa not affected by Cu-MOF.Moreover,RIgG@Cu-MOF has better biological and chemical stability than HRP-RIgG,and can be stored at room temperature for a long period.On this basis,by using Cu-MOF as a peroxidase mimic,RIgG@Cu-MOF can be used in colorimetric immunoassay.Benefiting from the ampilification effect of Cu-MOF,the detection limit(0.34 ng/mL)is three times lower than HRP-RIgG.Therefore,it is feasible to prepare novel enzyme-labeled antibodies based on MOF and apply them to colorimetric immunoassay.(2)Above results indicate that MOF is suitable for integrating with antibody,but its sensitivity is still limited by the poor catalytic activity and selectivity of Cu-MOF as an enzyme mimic.Accordingly,in this work,Zn-MOF was selected as a carrier to co-integrate with CEA antibody(anti-CEA)and alkaline phosphatase(ALP)to form a new dectection antibody,denoted as anti-CEA&ALP@Zn-MOF.The results show that this method can simultaneously realize the dual functions of enzyme catalysis and antibody specific recognition.On the one hand,compared with free ALP,zinc-MOF greatly improved activity and stability of ALP under extreme conditions such as high temperature and organic solvents can be achieved.On the other hand,it was demonstrated that the anti-CEA still remains its recognition ability and efficiency as free antibody.Based on this,anti-CEA&ALP@Zn-MOF can be used as a detection antibody to perform a colorimetric immunoassay of CEA antigen.Because of the cascade amplification effect of ALP and ascorbic acid,the detection limit of the sensor system can reach 22.7 pg/mL.(3)On the basis of above study,a cascade-ampified immunoassay was further developed by coupling glucose oxidase(GOx)and Fe-MOF with peroxidase-like activity.Such combination not only overcomes the shortcoming of Fe-MOF in poor selectivity,but also is beneficial to improve the overall activity.Upon the co-integration of prostate specific antigen antibody(anti-PSA)and GOx with Fe-MOF,a new detection antibody(anti-PSA&GOx@Fe-MOF)can be fabricated.Compared with free GOx and Fe-MOF systems,GOx@Fe-MOF not only has higher catalytic activity,but also exhibits improved biological and chemical stability due to the protective effect of Fe-MOF.More importantly,due to the confinement effect of Fe-MOF,the PSA antigen can be detected as low as 1.50 pg/mL.