A Hydrogen Bonded Non-Porous Organic-Inorganic Framework for Measuring Cysteine in Blood Plasma and Endogenous Cancer Cell.

An imbalance in cysteine (Cys) levels in the cells and plasma has been identified as the risk indicator for various human diseases. The structural similarity of cysteine with its congener homocysteine and glutathione offers challenges in its measurement. Herein, we report a hydrogen-bonded organic-inorganic framework of Cu(II) (HOIF) for the selective detection of cysteine over other biothiols. The non-fluorescent HOIF showed 12-fold green emission in the presence of cysteine. The monomeric unit of HOIF is stabilized via intermolecular hydrogen bonds, resulting in a non-porous network structure. Non-interference from homocysteine, glutathione, and other competitive bio-analytes revealed explicit affinity of HOIF for cysteine. Fluorimetric titration showed a wide working concentration window (650 nM-800 µM) for measuring cysteine in an aqueous medium. The mechanistic investigation involving HRMS, EPR, and UV-vis spectroscopic studies revealed the decomplexation of HOIF with Cys, resulting in a fluorescence turn-on response from the luminescent ligand. Validation using a commercial dye, "Cysteine Green", confirmed the prospect of HOIF for early diagnostic purposes. Utilizing the fluorescence turn-on property of HOIF in the presence of cysteine, we measured cysteine quantitatively in the blood plasma samples. Bio-imaging of endogenous cysteine in cancer cells indicated the ability of HOIF to monitor the intracellular cysteine.

Ready-to-use vertical flow paper device for instrument-free room temperature reverse transcription.

Paper-based nucleic acid detection and diagnosis are currently gaining much interest in point-of-care (POC) applications. The major steps involved in any nucleic acid amplification testing (NAAT) based diagnostics are nucleic acid isolation, reverse transcription (RT) (in the case of RNA), amplification and detection. RT is an important step in quantifying the viral load in case of disease diagnosis as well as quantifying gene expression levels in other molecular studies. cDNA synthesis is routinely carried out using a thermal cycler, with the process requiring temperatures between 40ºC to 65ºC. Here we report for the first time an instrument-free RT, performed at room temperature on cellulose-based paper devices. cDNA synthesis on paper was confirmed by RT-PCR and Sanger sequencing of the PCR products. Purified RNA from varied sources such as cell lysate, tissue and blood were used to test the methodology. Synthetic hepatitis C virus (HCV) RNA and human blood RNA were used as proof-of-concept to demonstrate the use of these devices in diagnostic applications. Further, ready-to-use paper-based reverse transcription (PRT) devices have been developed, wherein only the RNA sample is added on the device and the cDNA can be eluted after 30 min of incubation at room temperature. The devices were found to be stable for 30 days at - 20ºC storage. The cellulose-based PRT devices are simple, time saving and user-friendly for a complete instrument-free cDNA synthesis at room temperature.