Quantum Dots and Gold Nanoparticles as Scaffolds for Enzymatic Enhancement: Recent Advances and the Influence of Nanoparticle Size

Nanoparticle scaffolds can impart multiple benefits onto immobilized enzymes including enhanced stability, activity, and recoverability. The magnitude of these benefits is modulated by features inherent to the scaffold–enzyme conjugate, amongst which the size of the nanoscaffold itself can be critically important. In this review, we highlight the benefits of enzyme immobilization on nanoparticles and the factors affecting these benefits using quantum dots and gold nanoparticles as representative materials due to their maturity. We then review recent literature on the use of these scaffolds for enzyme immobilization and as a means to dissect the underlying mechanisms. Detailed analysis of the literature suggests that there is a “sweet-spot” for scaffold size and the ratio of immobilized enzyme to scaffold, with smaller scaffolds and lower enzyme:scaffold ratios generally providing higher enzymatic activities. We anticipate that ongoing studies of enzyme immobilization onto nanoscale scaffolds will continue to sharpen our understanding of what gives rise to beneficial characteristics and allow for the next important step, namely, that of translation to large-scale processes that exploit these properties.

Fabrication of dual catalytic microcapsules by mesoporous graphitic carbon nitride (mpg-C3N4) nanoparticle–enzyme conjugate stabilized emulsions

Dual catalytic microcapsules (MCs) were fabricated by simultaneous self-assembly and cross-linking of mpg-C3N4 nanoparticles (NPs) and lipase conjugates at oil–water interface.

An alternative Diagnostic design for Chronic Kidney Disease detection Based on Cystatin C

Objective: Chronic Kidney Disease (CKD) is usually diagnosed by measuring the glomerular filtration rate (GFR) and diagnostics are still inadequate at the clinical level. Most of the diagnostic kits available estimate GFR rate by determining clearance of serum creatinine using various instrumentation generally available at hospitals. Serum creatinine is considered the major marker for renal deficiency disorders. Additionally, it is also an indicator of muscle mass and dietary intake. Hence, the need for a more reliable marker for CKD arises. A low molecular weight protein cystatin C has been found to be a reliable biomarker for detection of kidney function as it is solely filtered by the glomerulus and not secreted by renal tubules.Method: The basic set up of the kit was designed using a syringe containing multi walled carbon nanotube (MWCNT) conjugated protease. Casein beads were immersed in PBS buffer in the syringe. The Glass/MWCNT/papain solid support was subsequently inserted into the syringe in such a way, that the beads came in contact with the immobilized enzyme conjugate. The inhibitory action of cystatin C against protease forms the basis for the functioning of the kitResults: Results indicated that papain while immobilization needs to be in dynamic conformation. At 37 C gave better activity as compared to protein immobilized at 4C. FTIR observations confirmed the physical adsorption on the MWCNTs. The experimentation confirmed the feasibility of using prototype for detection of cystatin C.Discussion: Papain conjugated with MWCNT indicated its temperature and pH stability. The initial design of the diagnostic kit for the detection of CKD has shown to be successful with a good detection range corresponding to stage I and II of CKD. Further testing needs to be done for the prototype using patient samples.Keywords: Chronic kidney disease, Diagnostic kit, Immobilized papain, Protease inhibitor.

Effect of Spacer and the Enzyme-Linked Immunosorbent Assay

The effect of spacers and the enzyme-linked immunosorbent assay (ELISA) formats on the functional parameters of assays such as lower detection limit, inhibitory concentration at 50 per cent (IC50), and specificity were studied. Enzyme conjugates having hydrophobic and hydrophilic spacers were prepared using O-isopropyl methylphosphonic acid (IMPA) and horseradish peroxidase (HRP) as an enzyme label. Comparison was made with reference to enzyme conjugate without any spacer. The present investigation revealed that the presence of a hydrophilic spacer in the enzyme conjugate significantly improves the sensitivity of assays. An enhanced IC50 value achieved was 0.01 μg mL−1 for free antigen detection by direct immunoassay using hydrophilic spacers and precoating of ELISA plates by secondary antibody. The use of a hydrophilic spacer might have helped in projecting the hapten in the aqueous phase, leading to enhanced antibody binding signal and improved sensitivity of the assay.