Detection of mercury and discrimination of breast cancerous and non-cancerous cells

Mr. Subhajit Chakraborty’s interview with Bio Patrika hosting “Vigyaan Patrika”, a series of author interviews. Subhajit is from West Bengal, India. He completed his B.Sc. in Chemistry from Ramakrishna Mission Vivekananda Centenary College, Rahara (2013). He did M.Sc. from the Presidency University, Kolkata in 2015. He joined the Ph.D. program (2016) in the Department of Chemistry of the Indian Institute of Science Education and Research Bhopal, under the supervision of Professor Saptarshi Mukherjee. His research work is mainly focused on the Fluorescence spectroscopy and development of luminescent metal nanomaterials to study their characteristic photophysical and morphological aspects and their applications in the various interdisciplinary field of sciences. Here, Subhajit talks about his work on ‘Protein-Templated Gold Nanoclusters as Specific Bio-Imaging Probe for the Detection of Hg(II) Ions in In Vivo and In Vitro Systems: Discriminating MDA-MB-231 and MCF10A Cells’ published in Analyst.

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How would you explain your paper’s key results to the non-scientific community?

Mercury (Hg) is a global poison for ecological systems particularly humans. Artisanal and small-scale gold mining (ASGM) is one of the main sources of mercury pollution after anthropogenic mercury emissions. The most dangerous aspect is that it can affect the central nervous system along with triggering other problems like hyper-irritability, fatigue, behavioral changes, hallucinations, etc. Mercury in its ionic form can readily combine with base pairs of DNA causing maximum damage to the endothelial cells, and cardiovascular functions. Additionally, it is expected that mercury will bind with thiol-containing biomolecules when present inside the body and can damper the normal metabolic activities of these bio-molecules. Thus, the highly sensitive detection of Hg(II) is very crucial (very low Limit of Detection (LOD)) and always a challenging task. Fluorophores that have prominent selective detection ability, have robust nature towards external physical and chemical perturbations, and can be used for single molecular investigations are termed as good sensors. They also have optimum quantum yields as well as very good photostability. Noble metal nanoclusters (NCs) templated by protein motifs have gained more impact of late as they render additional stability to the cluster core and thus generate interesting optical properties.

Schematic

Highlighting these characteristics, we have carried out the facile and optimized synthesis of gold nanocluster (AuNCs) within a protein (human serum albumin, HSA) template. Most interestingly, these AuNCs possess remarkable photo-, thermal- and core-cluster stability for more than a year (unaltered luminescence properties). We have characterized as-synthesized AuNCs using various spectroscopic and microscopic experiments. The core of these AuNCs consists of 25 gold atoms, which have been estimated by both the experimental and theoretical approaches. Importantly, we conclusively demonstrated the application of AuNCs as robust and photo-stable single molecular probes with the capability to function as highly sensitive (sub-nanomolar detection limit) and selective Hg(II) sensors. Our NCs proved to be quite beneficial in detecting the crucial and toxic Hg ions in solution as well as within a biomolecule. By artificially fabricating an Insulin-Hg complex, our AuNCs were able to efficiently estimate the bound Hg(II) under physiological conditions, thus emphasizing that the detection ability was conserved even at the single molecular resolution. Further study reveals that our AuNCs are also specific for intercellular localization. It specifically endocytoses inside the cancerous cell lines (MDA-MB-231) as revealed by the cell imaging study. Another important feature of this study is that these AuNCs also have the ability to detect Hg(II) ions inside the cell. The fluorescence of AuNCs was quenched in presence of Hg(II) ion inside the MDA-MB-231 cell lines. Thus, the output of our study apprises the sensitive in vivo as well as in vitro detection of Hg(II) ions using AuNCs as a probe.

The interesting feature of this work is that our AuNCs can detect the Hg(II) ions when it is bound with insulin biomolecule.

What are the possible consequences of these findings for your research area?

In this investigation, we have successfully developed intense red luminescent highly stable gold nanoclusters (AuNCs). These AuNCs demonstrated very efficient detection ability towards the Hg(II) ions by quenching their luminescence emission intensity. It is expected that the alteration and modification of the (S-S) bonds of insulin can occur in presence of Hg(II) ions. Our single molecular spectroscopic studies reveal that this detection limit can be extended up to the sub-nanomolar regime. The FCS analysis helped us to achieve very low LOD for Hg(II) detection (~0.01 nM) which is appreciably lower than the international standards set for Hg(II) levels in drinking water (10 nM) according to the U.S. EPA. The interesting feature of this work is that our AuNCs can detect the Hg(II) ions when it is bound with insulin biomolecule. This report shows another important feature of this investigation that is the specificity towards encapsulation of these AuNCs inside the triple-negative breast cancer (TNBC) cell, MDA-MB-231 in comparison to normal cell lines, MCF10A, and ability of detection of Hg(II) ions inside the MDA-MB-231 cell lines.

In a brief, this report can provide a very stable nanomaterial that can sensitively detect Hg(II) in in vitro as well as in vivo environments.

What was the exciting moment (eureka moment) during your research?

The overall journey of this work was exciting to me as each new experiment led to a new set of data, which in turn made the overall study more interesting and application-oriented. The preparation of the AuNCs and the observation of its intense luminescence had generated the initial excitement. The data quality, analysis, and correlation of all experimental techniques are always very satisfying at the end of the day for any researcher and I was no exception. Additionally, the concept that our work can be extended for in vivo cell imaging thereby sensing of Hg(II) inside the cell lines added color to the excitement.

What do you hope to do next?

As these AuNCs can detect Hg(II) ions in various systems, we can use this to detect mercury in drinking water and can thereby analyze the quality of the water. In another aspect, the biomolecular dynamics inside the cell are always important to investigate. We are planning to use these AuNCs as a bio-marker and will try to investigate the inter-cellular environments. We shall also monitor the dynamics of various bio-molecules inside the cells by utilizing FCS and FLIM analyses.

Where do you seek scientific inspiration?

I always feel investigating the unrevealed facts and knowing the unknown inspires me a lot. The research environment in my lab and more importantly, the inspiration from my thesis supervisor Professor Saptarshi Mukherjee all the time (and even today) helps me to move ahead every day. New research reports in various journals, handling new instruments, analysis of new data, etc. definitely creates interest in me and motivates me to work and explore new(er) aspects of science.

How do you intend to help Indian science improve?

Indian science has already evolved to a great extent, still more improvements are needed. According to me, the interest in science and intention of scientific investigation can be created from the school level of education. Another important factor is the development of genuine interest in the understanding of new knowledge inside and outside the laboratory. If I get a chance, then I would like to focus on my further research by creating a group to understand in-depth fundamental aspects of the science along with the results which can be applied to our society. There should be a balance between fundamental and applied research. We need good teachers at all strata so that the thirst among students always remains and they can ask and address the correct questions which will take care of the societal needs. Today Indian science needs a proper synergy between education and research.

Reference

S. Chakraborty, A. Nandy, S. Ghosh, N. K. Das, S. Parveen, S. Datta and S. Mukherjee, Protein-Templated Gold Nanoclusters as Specific Bio-Imaging Probe for the Detection of Hg(II) Ions in In Vivo and In Vitro Systems: Discriminating MDA-MB-231 and MCF10A Cells. Analyst 2021, 146, 1455-1463.

Email: subhajit16@iiserb.ac.in, saptarshi@iiserb.ac.in

Edited by: Ritvi Shah

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