The Royal Society of Chemistry eBooks, Oct 7, 2016
The in vivo mammalian alkaline Comet assay is used for assessing the DNA-damaging potential of ch... more The in vivo mammalian alkaline Comet assay is used for assessing the DNA-damaging potential of chemicals in cells and tissues of rats and mice. The assay has been adopted by the OECD, as a second-tier test, in the standard battery of genotoxicity tests. This chapter outlines the protocol followed for performing the in vivo Comet assay in cells of blood and different organs e.g. liver, kidney, brain, spleen and bone marrow of mouse.
Titanium dioxide nanoparticles (TiO(2) NPs), widely used in consumer products, paints, pharmaceut... more Titanium dioxide nanoparticles (TiO(2) NPs), widely used in consumer products, paints, pharmaceutical preparations and so on, have been shown to induce cytotoxicity, genotoxicity and carcinogenic responses in vitro and in vivo. The present study revealed that TiO(2) NPs induce significant (p < 0.05) oxidative DNA damage by the Fpg-Comet assay even at 1 µg/ml concentration. A corresponding increase in the micronucleus frequency was also observed. This could be attributed to the reduced glutathione levels with concomitant increase in lipid peroxidation and reactive oxygen species generation. Furthermore, immunoblot analysis revealed an increased expression of p53, BAX, Cyto-c, Apaf-1, caspase-9 and caspase-3 and decreased the level of Bcl-2 thereby indicating that apoptosis induced by TiO(2) NPs occurs via the caspase-dependent pathway. This study systematically shows that TiO(2) NPs induce DNA damage and cause apoptosis in HepG2 cells even at very low concentrations. Hence the use of such nanoparticles should be carefully monitored.
Nanoparticles possess size-dependent chemical and physical characteristics that enable interestin... more Nanoparticles possess size-dependent chemical and physical characteristics that enable interesting and correlated approaches for dealing with fundamental biological molecules. Despite the significant development of nanoscience, interactions of nanoscale objects with living systems is less known. When a nanoparticle (NP) encounters a biological fluid, biomolecules spontaneously form adsorption layers around the NP, called a “protein corona” (PC). The corona's composition depends on the time-dependent environmental conditions, which determines the NP's fate within living organisms. The PC consists of two poorly delimited layers, known as the “hard corona” (HC) and “soft corona” (SC), which is affected by the complexity of the environment and the protein–surface equilibrium formed during in vivo blood circulation. This chapter is focused on the investigation of the corona formation of adsorbed proteins around nanoparticles depending on the type of characterization technique. Protein corona–NP complexes are further characterized by integrating information on morphology and also on the structure/composition of the PC. Thus, multi-disciplinary approaches are highlighted in order to obtain much more information about the PC and its properties to fully understand the real impact of the PC on nanoparticles' surface and their various therapeutics applications.
Zinc oxide nanoparticles (ZnO NPs) with their wide range of consumer applications in day-to-day l... more Zinc oxide nanoparticles (ZnO NPs) with their wide range of consumer applications in day-to-day life received great attention to evaluate their effects in humans. This study has been attempted to elucidate the DNA damage response mechanism in a dermal model exposed to ZnO NPs through Ataxia Telangiectasia Mutated (ATM)-mediated ChK1-dependent G2/M arrest. Further, viability parameters and mechanism involved in the cell death with special reference to the consequences arising due to DNA damage were explored. Our study showed that ZnO NPs at concentrations 5 and 10 µg/ml induced significant cytotoxic effect in skin cell line. Moreover, the results confirmed generation of reactive oxygen species (ROS) induces the cell death by genotoxic insult, leading to mitochondrial membrane depolarisation and cell cycle arrest. Subsequently, ZnO NPs treatment created DNA damage as confirmed via Comet assay (increase in olive tail moment), micronucleus assay (increase in micronucleus formation), dou...
Bioinspired, Biomimetic and Nanobiomaterials, 2016
This paper reports the impact of bovine serum albumin (BSA) coating on gold (Au) nanoparticles (N... more This paper reports the impact of bovine serum albumin (BSA) coating on gold (Au) nanoparticles (NPs) with sizes of 15, 30, 50 and 70 nm on cellular uptake and haemolysis of human red blood cells (RBCs). BSA coating on gold NPs imparts extra stability in high-glutathione-containing medium, which is a major prerequisite for NPs being developed for delivery applications. BSA coating on gold NPs was characterised by Fourier transform infrared spectroscopy, whereas cellular uptake was estimated by ultraviolet–visible spectrophotometry and flow cytometry. The cellular uptake results show that the internalisation of bare gold NPs is size dependent; however, upon BSA conjugation, uptake becomes independent of particle size. Cytocompatibility of bare and BSA-coated gold NPs was assessed by MTT assay, a common method to evaluate the biocompatibility of nanomaterials, and found non-toxic. However, when bare gold NPs were exposed to human RBCs, the NPs exerted significant haemolysis, which sugg...
Precision in experiment is one of the most important parameter in the field of Science. To obtain... more Precision in experiment is one of the most important parameter in the field of Science. To obtain the precise result sophisticated instrumentations are required. In the field of forensic science, a forensic scientist must be proficient in assimilating knowledge and skills to examine, analyze, interpret, reporting, and give expert opinion in support of evidence. These aforementioned skills can work only when examination will perform with precision. Nowadays, in criminal investigations forensic experts are using biotechnology based methods for sample analysis. Basically, Forensic investigation is the application of scientific tools and specific scientific facts that can help to solve the legal problems. The best example of application of biotechnology in Forensic science is DNA Forensic. This branch of forensic science is now transforming many aspects of criminal investigation. By the help of biotechnology methods, analysis of DNA samples allows precise identifications to be made from very minute evidence collected at the scene of crime.This application of biotechnology in forensic investigation emerged a new branch of forensic science known as “Forensic Biotechnology”. Typically, forensic biotechnology is linked with DNA fingerprinting. By identifying DNA recovered from the biological evidences such as biological fluids, hair or tissues collected at the scene of crime, a forensic biotechnologist can link to a suspect to the crime scene or identify an unknown individual. Another application of forensic biotechnology is its use to identify and monitor non-human organisms, such as endangered species, through DNA fingerprinting. Application of Flow Cytometry in Forensic Medicine
Microfluidics is the technology or system wherein the behavior of fluids' is studied onto a m... more Microfluidics is the technology or system wherein the behavior of fluids' is studied onto a miniaturized device composed of chambers and tunnels. In biological and biomedical sciences, microfluidic technology/system or device serves as an ultra-high-output approach capable of detecting and separating the biomolecules present even in trace quantities. Given the essential role of protein, the identification and quantification of proteins help understand the various living systems' biological function regulation. Microfluidics has enormous potential to enable biological investigation at the cellular and molecular level and maybe a fair substitution of the sophisticated instruments/equipment used for proteomics, genomics, and metabolomics analysis. The current advancement in microfluidic systems' development is achieving momentum and opening new avenues in developing innovative and hybrid methodologies/technologies. This chapter attempts to expound the micro/nanofluidic systems/devices for their wide-ranging application to detect and separate protein. It covers microfluidic chip electrophoresis, microchip gel electrophoresis, and nanofluidic systems as protein separation systems, while methods such as spectrophotometric, mass spectrometry, electrochemical detection, magneto-resistive sensors and dynamic light scattering (DLS) are discussed as proteins' detection system.
Jpc – Journal Of Planar Chromatography – Modern Tlc, Aug 1, 2020
Food adulteration is a primary global concern for public health, especially in developing countri... more Food adulteration is a primary global concern for public health, especially in developing countries, due to the lack of monitoring and appropriate policy developments and implementations. Synthetic colors and dyes are used to enhance the appearance of food products. Metanil Yellow (MY), a non-permitted, toxic, chief additive adulterant is majorly used in sweets, pulses, and turmeric in India owing to its color that ranges from yellow to orange. Owing to this, we have assessed the presence of MY in sweets like “jalebi” and “laddu” from branded and local vendors. We present a method to isolate the additive from the sweets with the help of acetic acid, followed by screening through colorimetric assay. The chemical characterization of MY was conducted using a multi-plate ultraviolet–visible (UV–Vis) spectrophotometer and attenuated total reflectance–Fourier transform infrared spectroscopy (ATR–FTIR). The appearance of magenta/pink color indicates the presence of MY in the chemical spot test. Also, thin-layer chromatography (TLC) was applied to validate the presence of this adulterant by comparing the peaks of reference (MY) with the extracted samples. TLC of food color extracted from the samples is an easy, efficient, and cost-effective method to detect toxic adulterants like MY. Our study demonstrates a simple food color extraction procedure, with easy and sensitive chemical assays, for the detection of MY in sweets. It also offers reasonable sensitivity and can be used to screen large numbers of samples in a short time.
The Royal Society of Chemistry eBooks, Oct 7, 2016
The in vivo mammalian alkaline Comet assay is used for assessing the DNA-damaging potential of ch... more The in vivo mammalian alkaline Comet assay is used for assessing the DNA-damaging potential of chemicals in cells and tissues of rats and mice. The assay has been adopted by the OECD, as a second-tier test, in the standard battery of genotoxicity tests. This chapter outlines the protocol followed for performing the in vivo Comet assay in cells of blood and different organs e.g. liver, kidney, brain, spleen and bone marrow of mouse.
Titanium dioxide nanoparticles (TiO(2) NPs), widely used in consumer products, paints, pharmaceut... more Titanium dioxide nanoparticles (TiO(2) NPs), widely used in consumer products, paints, pharmaceutical preparations and so on, have been shown to induce cytotoxicity, genotoxicity and carcinogenic responses in vitro and in vivo. The present study revealed that TiO(2) NPs induce significant (p < 0.05) oxidative DNA damage by the Fpg-Comet assay even at 1 µg/ml concentration. A corresponding increase in the micronucleus frequency was also observed. This could be attributed to the reduced glutathione levels with concomitant increase in lipid peroxidation and reactive oxygen species generation. Furthermore, immunoblot analysis revealed an increased expression of p53, BAX, Cyto-c, Apaf-1, caspase-9 and caspase-3 and decreased the level of Bcl-2 thereby indicating that apoptosis induced by TiO(2) NPs occurs via the caspase-dependent pathway. This study systematically shows that TiO(2) NPs induce DNA damage and cause apoptosis in HepG2 cells even at very low concentrations. Hence the use of such nanoparticles should be carefully monitored.
Nanoparticles possess size-dependent chemical and physical characteristics that enable interestin... more Nanoparticles possess size-dependent chemical and physical characteristics that enable interesting and correlated approaches for dealing with fundamental biological molecules. Despite the significant development of nanoscience, interactions of nanoscale objects with living systems is less known. When a nanoparticle (NP) encounters a biological fluid, biomolecules spontaneously form adsorption layers around the NP, called a “protein corona” (PC). The corona's composition depends on the time-dependent environmental conditions, which determines the NP's fate within living organisms. The PC consists of two poorly delimited layers, known as the “hard corona” (HC) and “soft corona” (SC), which is affected by the complexity of the environment and the protein–surface equilibrium formed during in vivo blood circulation. This chapter is focused on the investigation of the corona formation of adsorbed proteins around nanoparticles depending on the type of characterization technique. Protein corona–NP complexes are further characterized by integrating information on morphology and also on the structure/composition of the PC. Thus, multi-disciplinary approaches are highlighted in order to obtain much more information about the PC and its properties to fully understand the real impact of the PC on nanoparticles' surface and their various therapeutics applications.
Zinc oxide nanoparticles (ZnO NPs) with their wide range of consumer applications in day-to-day l... more Zinc oxide nanoparticles (ZnO NPs) with their wide range of consumer applications in day-to-day life received great attention to evaluate their effects in humans. This study has been attempted to elucidate the DNA damage response mechanism in a dermal model exposed to ZnO NPs through Ataxia Telangiectasia Mutated (ATM)-mediated ChK1-dependent G2/M arrest. Further, viability parameters and mechanism involved in the cell death with special reference to the consequences arising due to DNA damage were explored. Our study showed that ZnO NPs at concentrations 5 and 10 µg/ml induced significant cytotoxic effect in skin cell line. Moreover, the results confirmed generation of reactive oxygen species (ROS) induces the cell death by genotoxic insult, leading to mitochondrial membrane depolarisation and cell cycle arrest. Subsequently, ZnO NPs treatment created DNA damage as confirmed via Comet assay (increase in olive tail moment), micronucleus assay (increase in micronucleus formation), dou...
Bioinspired, Biomimetic and Nanobiomaterials, 2016
This paper reports the impact of bovine serum albumin (BSA) coating on gold (Au) nanoparticles (N... more This paper reports the impact of bovine serum albumin (BSA) coating on gold (Au) nanoparticles (NPs) with sizes of 15, 30, 50 and 70 nm on cellular uptake and haemolysis of human red blood cells (RBCs). BSA coating on gold NPs imparts extra stability in high-glutathione-containing medium, which is a major prerequisite for NPs being developed for delivery applications. BSA coating on gold NPs was characterised by Fourier transform infrared spectroscopy, whereas cellular uptake was estimated by ultraviolet–visible spectrophotometry and flow cytometry. The cellular uptake results show that the internalisation of bare gold NPs is size dependent; however, upon BSA conjugation, uptake becomes independent of particle size. Cytocompatibility of bare and BSA-coated gold NPs was assessed by MTT assay, a common method to evaluate the biocompatibility of nanomaterials, and found non-toxic. However, when bare gold NPs were exposed to human RBCs, the NPs exerted significant haemolysis, which sugg...
Precision in experiment is one of the most important parameter in the field of Science. To obtain... more Precision in experiment is one of the most important parameter in the field of Science. To obtain the precise result sophisticated instrumentations are required. In the field of forensic science, a forensic scientist must be proficient in assimilating knowledge and skills to examine, analyze, interpret, reporting, and give expert opinion in support of evidence. These aforementioned skills can work only when examination will perform with precision. Nowadays, in criminal investigations forensic experts are using biotechnology based methods for sample analysis. Basically, Forensic investigation is the application of scientific tools and specific scientific facts that can help to solve the legal problems. The best example of application of biotechnology in Forensic science is DNA Forensic. This branch of forensic science is now transforming many aspects of criminal investigation. By the help of biotechnology methods, analysis of DNA samples allows precise identifications to be made from very minute evidence collected at the scene of crime.This application of biotechnology in forensic investigation emerged a new branch of forensic science known as “Forensic Biotechnology”. Typically, forensic biotechnology is linked with DNA fingerprinting. By identifying DNA recovered from the biological evidences such as biological fluids, hair or tissues collected at the scene of crime, a forensic biotechnologist can link to a suspect to the crime scene or identify an unknown individual. Another application of forensic biotechnology is its use to identify and monitor non-human organisms, such as endangered species, through DNA fingerprinting. Application of Flow Cytometry in Forensic Medicine
Microfluidics is the technology or system wherein the behavior of fluids' is studied onto a m... more Microfluidics is the technology or system wherein the behavior of fluids' is studied onto a miniaturized device composed of chambers and tunnels. In biological and biomedical sciences, microfluidic technology/system or device serves as an ultra-high-output approach capable of detecting and separating the biomolecules present even in trace quantities. Given the essential role of protein, the identification and quantification of proteins help understand the various living systems' biological function regulation. Microfluidics has enormous potential to enable biological investigation at the cellular and molecular level and maybe a fair substitution of the sophisticated instruments/equipment used for proteomics, genomics, and metabolomics analysis. The current advancement in microfluidic systems' development is achieving momentum and opening new avenues in developing innovative and hybrid methodologies/technologies. This chapter attempts to expound the micro/nanofluidic systems/devices for their wide-ranging application to detect and separate protein. It covers microfluidic chip electrophoresis, microchip gel electrophoresis, and nanofluidic systems as protein separation systems, while methods such as spectrophotometric, mass spectrometry, electrochemical detection, magneto-resistive sensors and dynamic light scattering (DLS) are discussed as proteins' detection system.
Jpc – Journal Of Planar Chromatography – Modern Tlc, Aug 1, 2020
Food adulteration is a primary global concern for public health, especially in developing countri... more Food adulteration is a primary global concern for public health, especially in developing countries, due to the lack of monitoring and appropriate policy developments and implementations. Synthetic colors and dyes are used to enhance the appearance of food products. Metanil Yellow (MY), a non-permitted, toxic, chief additive adulterant is majorly used in sweets, pulses, and turmeric in India owing to its color that ranges from yellow to orange. Owing to this, we have assessed the presence of MY in sweets like “jalebi” and “laddu” from branded and local vendors. We present a method to isolate the additive from the sweets with the help of acetic acid, followed by screening through colorimetric assay. The chemical characterization of MY was conducted using a multi-plate ultraviolet–visible (UV–Vis) spectrophotometer and attenuated total reflectance–Fourier transform infrared spectroscopy (ATR–FTIR). The appearance of magenta/pink color indicates the presence of MY in the chemical spot test. Also, thin-layer chromatography (TLC) was applied to validate the presence of this adulterant by comparing the peaks of reference (MY) with the extracted samples. TLC of food color extracted from the samples is an easy, efficient, and cost-effective method to detect toxic adulterants like MY. Our study demonstrates a simple food color extraction procedure, with easy and sensitive chemical assays, for the detection of MY in sweets. It also offers reasonable sensitivity and can be used to screen large numbers of samples in a short time.
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