Displaying publications 81 - 100 of 3431 in total

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  1. Development of multiplex loop mediated isothermal amplification (m-LAMP) label-based gold nanoparticles lateral flow dipstick biosensor for detection of pathogenic Leptospira
    Nurul Najian AB, Engku Nur Syafirah EA, Ismail N, Mohamed M, Yean CY
    Anal Chim Acta, 2016 Jan 15;903:142-8.
    PMID: 26709307 DOI: 10.1016/j.aca.2015.11.015
    In recent years extensive numbers of molecular diagnostic methods have been developed to meet the need of point-of-care devices. Efforts have been made towards producing rapid, simple and inexpensive DNA tests, especially in the diagnostics field. We report on the development of a label-based lateral flow dipstick for the rapid and simple detection of multiplex loop-mediated isothermal amplification (m-LAMP) amplicons. A label-based m-LAMP lateral flow dipstick assay was developed for the simultaneous detection of target DNA template and a LAMP internal control. This biosensor operates through a label based system, in which probe-hybridization and the additional incubation step are eliminated. We demonstrated this m-LAMP assay by detecting pathogenic Leptospira, which causes the re-emerging disease Leptospirosis. The lateral flow dipstick was developed to detect of three targets, the LAMP target amplicon, the LAMP internal control amplicon and a chromatography control. Three lines appeared on the dipstick, indicating positive results for all representative pathogenic Leptospira species, whereas two lines appeared, indicating negative results, for other bacterial species. The specificity of this biosensor assay was 100% when it was tested with 13 representative pathogenic Leptospira species, 2 intermediate Leptospira species, 1 non-pathogenic Leptospira species and 28 other bacteria species. This study found that this DNA biosensor was able to detect DNA at concentrations as low as 3.95 × 10(-1) genomic equivalent ml(-1). An integrated m-LAMP and label-based lateral flow dipstick was successfully developed, promising simple and rapid visual detection in clinical diagnostics and serving as a point-of-care device.
    Matched MeSH terms: DNA
  2. Target Induced-DNA strand displacement reaction using gold nanoparticle labeling for hepatitis E virus detection
    Ngamdee T, Yin LS, Vongpunsawad S, Poovorawan Y, Surareungchai W, Lertanantawong B
    Anal Chim Acta, 2020 Oct 16;1134:10-17.
    PMID: 33059855 DOI: 10.1016/j.aca.2020.08.018
    DNA strand displacement is an attractive, enzyme-free target hybridization strategy for nano-biosensing. The target DNA induces a strand displacement reaction by replacing the pre-hybridized strand that is labeled with gold nanoparticles (AuNPs). Thus, the amount of displaced-AuNP-labeled strand is proportional to the amount of target DNA in the sample. The use of a magnetogenosensing technique to isolate the target DNA allows for a simple, one-pot detection approach, which minimizes possible carry-over contamination and pipetting errors. We sought a proof-of-concept for this technology in its ability to detect DNA-equivalent of hepatitis E virus (HEV), which causes acute viral hepatitis for which rapid and simple diagnostic methods remain limited. Signal detection was done via visual observation, spectrophotometry, and electrochemistry. The sensor demonstrated good sensitivity with detection limits of 10 pM (visual), 10 pM (spectrophotometry) and 1 fM (electrochemical). This sensor also exhibited high specificity for real target amplicons and could discriminate between perfect and mismatched sequences. Lyophilized biosensor reagents stored at 4 °C, 25 °C, and outdoor ambient temperature, were stable for up to 90, 50, and 40 days, respectively. The integration of magnetic separation and target DNA-induced strand displacement reaction in a dry reagent form makes the sensing platform easy-to-use and suitable for field settings.
    Matched MeSH terms: DNA
  3. Development of a thermostabilised triplex LAMP assay with dry-reagent four target lateral flow dipstick for detection of Entamoeba histolytica and non-pathogenic Entamoeba spp
    Foo PC, Chan YY, Mohamed M, Wong WK, Nurul Najian AB, Lim BH
    Anal Chim Acta, 2017 May 08;966:71-80.
    PMID: 28372729 DOI: 10.1016/j.aca.2017.02.019
    This study highlighted the development of a four target nitrocellulose-based nucleic acid lateral flow immunoassay biosensor in a dry-reagent strip format for interpretation of double-labelled double-stranded amplicons from thermostabilised triplex loop-mediated isothermal amplification assay. The DNA biosensor contained two test lines which captured biotin and texas red labelled amplicons; a LAMP internal amplification control line that captured digoxigenin labelled amplicon; and a chromatography control line that validated the functionality of the conjugated gold nanoparticles and membrane. The red lines on detection pad were generated when the gold nanoparticles conjugated antibody bound to the fluorescein labelled amplicons, and the capture agents bound to their specific hapten on the other 5' end of the double-stranded amplicon. The applicability of this DNA biosensor was demonstrated using amoebiasis-causing Entamoeba histolytica simultaneously with the non-pathogenic but morphologically identical Entamoeba dispar and Entamoeba moshkovskii. The biosensor detection limit was 10 E. histolytica trophozoites, and revealed 100% specificity when it was evaluated against 3 medically important Entamoeba species and 75 other pathogenic microorganisms. Heat stability test showed that the biosensor was stable for at least 181 days at ambient temperature. This ready-to-use and cold-chain-free biosensor facilitated the post-LAMP analysis based on visualisation of lines on strip instead of observation of amplicon patterns in agarose gel.
    Matched MeSH terms: DNA, Protozoan/analysis
  4. Electrochemical DNA detection of hepatitis E virus genotype 3 using PbS quantum dot labelling
    Ngo DB, Chaibun T, Yin LS, Lertanantawong B, Surareungchai W
    Anal Bioanal Chem, 2021 Feb;413(4):1027-1037.
    PMID: 33236225 DOI: 10.1007/s00216-020-03061-1
    The aim of this study was to develop a highly specific electrochemical DNA sensor using functionalized lead sulphide (PbS) quantum dots for hepatitis E virus genotype 3 (HEV3) DNA target detection. Functionalized-PbS quantum dots (QDs) were used as an electrochemical label for the detection of HEV3-DNA target by the technique of square wave anodic stripping voltammetry (SWASV). The functionalized-PbS quantum dots were characterized by UV-vis, FTIR, XRD, TEM and zeta potential techniques. As-prepared, functionalized-PbS quantum dots have an average size of 4.15 ± 1.35 nm. The detection platform exhibited LOD and LOQ values of 1.23 fM and 2.11 fM, respectively. HEV3-DNA target spiked serum is also reported.Graphical abstract.
    Matched MeSH terms: DNA, Viral/blood*; DNA, Viral/genetics
  5. An ultrasensitive hollow-silica-based biosensor for pathogenic Escherichia coli DNA detection
    Ariffin EY, Lee YH, Futra D, Tan LL, Karim NHA, Ibrahim NNN, et al.
    Anal Bioanal Chem, 2018 Mar;410(9):2363-2375.
    PMID: 29504083 DOI: 10.1007/s00216-018-0893-1
    A novel electrochemical DNA biosensor for ultrasensitive and selective quantitation of Escherichia coli DNA based on aminated hollow silica spheres (HSiSs) has been successfully developed. The HSiSs were synthesized with facile sonication and heating techniques. The HSiSs have an inner and an outer surface for DNA immobilization sites after they have been functionalized with 3-aminopropyltriethoxysilane. From field emission scanning electron microscopy images, the presence of pores was confirmed in the functionalized HSiSs. Furthermore, Brunauer-Emmett-Teller (BET) analysis indicated that the HSiSs have four times more surface area than silica spheres that have no pores. These aminated HSiSs were deposited onto a screen-printed carbon paste electrode containing a layer of gold nanoparticles (AuNPs) to form a AuNP/HSiS hybrid sensor membrane matrix. Aminated DNA probes were grafted onto the AuNP/HSiS-modified screen-printed electrode via imine covalent bonds with use of glutaraldehyde cross-linker. The DNA hybridization reaction was studied by differential pulse voltammetry using an anthraquinone redox intercalator as the electroactive DNA hybridization label. The DNA biosensor demonstrated a linear response over a wide target sequence concentration range of 1.0×10-12-1.0×10-2 μM, with a low detection limit of 8.17×10-14 μM (R2 = 0.99). The improved performance of the DNA biosensor appeared to be due to the hollow structure and rough surface morphology of the hollow silica particles, which greatly increased the total binding surface area for high DNA loading capacity. The HSiSs also facilitated molecule diffusion through the silica hollow structure, and substantially improved the overall DNA hybridization assay. Graphical abstract Step-by-step DNA biosensor fabrication based on aminated hollow silica spheres.
    Matched MeSH terms: DNA, Bacterial/analysis*; DNA, Bacterial/genetics
  6. IQPA: isothermal nucleic acid amplification-based immunoassay using DNAzyme as the reporter system
    Loh Q, Omar N, Glökler J, Lim TS
    Anal Biochem, 2014 Oct 15;463:67-9.
    PMID: 24972268 DOI: 10.1016/j.ab.2014.06.012
    Immunoassays are often coupled to peroxidase activity for antigen detection. Sensitivity and speed of detection has been increased by the advent of hybrid methods such as immuno-PCR (polymerase chain reaction). However, a more simplified immunoassay that retains both colorimetric peroxidase detection and effective DNA amplification in a setting closer to field application conditions has been nonexistent. Here we describe a method that successfully combines a competitive immunoassay with the new isothermal quadruplex-primed amplification (QPA) to generate excess quadruplex reporter molecules with intrinsic peroxidase DNAzyme activity.
    Matched MeSH terms: DNA/analysis*; DNA/metabolism; DNA, Catalytic/metabolism*
  7. Phage N15 protelomerase resolves its tos recognition site into hairpin telomeres within mammalian cells
    Liew PS, Chen Q, Ng AWR, Chew YC, Ravin NV, Sim EUH, et al.
    Anal Biochem, 2019 10 15;583:113361.
    PMID: 31306622 DOI: 10.1016/j.ab.2019.113361
    Phage N15 protelomerase (TelN) cleaves double-stranded circular DNA containing a telomerase-occupancy-site (tos) and rejoins the resulting linear-ends to form closed-hairpin-telomeres in Escherichia coli (E. coli). Continued TelN expression is essential to support resolution of the linear structure. In mammalian cells, no enzyme with TelN-like activities has been found. In this work, we show that phage TelN, expressed transiently and stably in human and mouse cells, recapitulates its native activities in these exogenous environments. We found TelN to accurately resolve tos-DNA in vitro and in vivo within human and mouse cells into linear DNA-containing terminal telomeres that are resistant to RecBCD degradation, a hallmark of protelomerase processing. In stable cells, TelN activity was detectable for at least 60 days, which suggests the possibility of limited silencing of its expression. Correspondingly, linear plasmid containing a 100 kb human β-globin gene expressed for at least 120 h in non-β-globin-expressing mouse cells with TelN presence. Our results demonstrate TelN is able to cut and heal DNA as hairpin-telomeres within mammalian cells, providing a tool for creating novel structures by DNA resolution in these hosts. The TelN protelomerase may be useful for exploring novel technologies for genome interrogation and chromosome engineering.
    Matched MeSH terms: DNA/metabolism*; DNA Replication/physiology*
  8. Advancements in electrochemical DNA sensor for detection of human papilloma virus - A review
    Rasouli E, Shahnavaz Z, Basirun WJ, Rezayi M, Avan A, Ghayour-Mobarhan M, et al.
    Anal Biochem, 2018 09 01;556:136-144.
    PMID: 29981317 DOI: 10.1016/j.ab.2018.07.002
    Human papillomavirus (HPV) is one of the most common sexually transmitted disease, transmitted through intimate skin contact or mucosal membrane. The HPV virus consists of a double-stranded circular DNA and the role of HPV virus in cervical cancer has been studied extensively. Thus it is critical to develop rapid identification method for early detection of the virus. A portable biosensing device could give rapid and reliable results for the identification and quantitative determination of the virus. The fabrication of electrochemical biosensors is one of the current techniques utilized to achieve this aim. In such electrochemical biosensors, a single-strand DNA is immobilized onto an electrically conducting surface and the changes in electrical parameters due to the hybridization on the electrode surface are measured. This review covers the recent developments in electrochemical DNA biosensors for the detection of HPV virus. Due to the several advantages of electrochemical DNA biosensors, their applications have witnessed an increased interest and research focus nowadays.
    Matched MeSH terms: DNA, Single-Stranded/analysis*; DNA, Viral/analysis*
  9. DNA-templated silver nanocluster for live-intracellular FOXP3 detection
    Lee SY, Fazlina N, Tye GJ
    Anal Biochem, 2019 09 15;581:113352.
    PMID: 31260647 DOI: 10.1016/j.ab.2019.113352
    DNA-templated silver nanocluster (AgNC), a new promising fluorescence probe has gained importance in biosensing and bioimaging in recent years. We employed a label-free AgNC to detect an intracellular transcription factor known as forkhead box p3 (FOXP3), which is the master regulator of regulatory T cells (Tregs) suppressive function. We developed an optimized method for the detection of messenger ribonucleic acid (mRNA) of FOXP3 by hybridizing AgNC and G-rich to the target FOXP3 mRNA of a MCF-7 cells. MCF-7 cells are chosen as a model as it readily expresses FOXP3. The hybridized samples were examined with UV illuminator and further verified with fluorescence spectroscopy, fluorescence microscope and flow cytometry. The successful hybridization of a three-way junction with AgNC, G-rich and mRNA FOXP3 target generated an improved fluorescence intensity with a spectral shift. We have successfully delivered the green fluorescing AgNC and G-rich into MCF-7 cells, producing a shift to red fluorescing cells corroborated by flow cytometry results. In summary, our approach enables the detection of intracellular FOXP3 nucleic acid and holds considerable potential in establishing a non-lethal intracellular detection system which would be crucial for the isolation of regulatory T-cells (Tregs) when combined with other cell surface markers.
    Matched MeSH terms: DNA/chemistry*
  10. Molecularly imprinted polymers-based DNA biosensors
    Nawaz N, Abu Bakar NK, Muhammad Ekramul Mahmud HN, Jamaludin NS
    Anal Biochem, 2021 10 01;630:114328.
    PMID: 34363786 DOI: 10.1016/j.ab.2021.114328
    In multiple biological processes, molecular recognition performs an integral role in detecting bio analytes. Molecular imprinted polymers (MIPs) are tailored sensing materials that can biomimic the biologic ligands and can detect specific target molecules selectively and sensitively. The formulation of molecularly imprinted polymers is followed by the formulation of a control termed as non-imprinted polymer (NIP), which, in the absence of a template, is commonly formulated to evaluate whether distinctive imprints have been produced for the template. Given the difficulties confronting bioanalytical researchers, it is inevitable that this strategy would come out as a central route of multidisciplinary studies to create extremely promising stable artificial receptors as a replacement or accelerate biological matrices. The ease of synthesis, low cost, capability to 'tailor' recognition element for analyte molecules, and stability under harsh environments make MIPs promising candidates as a recognition tool for biosensing. Compared to biological systems, molecular imprinting techniques have several advantages, including high recognition ability, long-term durability, low cost, and robustness, allowing molecularly imprinted polymers to be employed in drug delivery, biosensor technology, and nanotechnology. Molecular imprinted polymer-based sensors still have certain shortcomings in determining biomacromolecules (nucleic acid, protein, lipids, and carbohydrates), considering the vast volume of the latest literature on biomicromolecules. These potential materials are still required to address a few weaknesses until gaining their position in recognition of biomacromolecules. This review aims to highlight the current progress in molecularly imprinted polymers (MIPs)-based sensors for the determination of deoxyribonucleic acid (DNA) or nucleobases.
    Matched MeSH terms: DNA/analysis*
  11. Large BACs transfect more efficiently in circular topology
    Wong YC, Osahor A, Al-Ajli FOM, Narayanan K
    Anal Biochem, 2021 10 01;630:114324.
    PMID: 34363787 DOI: 10.1016/j.ab.2021.114324
    The effect of DNA topology on transfection efficiency of mammalian cells has been widely tested on plasmids smaller than 10 kb, but little is known for larger DNA vectors carrying intact genomic DNA containing introns, exons, and regulatory regions. Here, we demonstrate that circular BACs transfect more efficiently than covalently closed linear BACs. We found up to 3.1- and 8.9- fold higher eGFP expression from circular 11 kb and 100 kb BACs, respectively, compared to linear BACs. These findings provide insights for improved vector development for gene delivery and expression studies of large intact transgenes in mammalian cells.
    Matched MeSH terms: DNA/genetics*
  12. Enzyme-linked amperometric electrochemical genosensor assay for the detection of PCR amplicons on a streptavidin-treated screen-printed carbon electrode
    Yean CY, Kamarudin B, Ozkan DA, Yin LS, Lalitha P, Ismail A, et al.
    Anal Chem, 2008 Apr 15;80(8):2774-9.
    PMID: 18311943 DOI: 10.1021/ac702333x
    A general purpose enzyme-based amperometric electrochemical genosensor assay was developed wherein polymerase chain reaction (PCR) amplicons labeled with both biotin and fluorescein were detected with peroxidase-conjugated antifluorescein antibody on a screen-printed carbon electrode (SPCE). As a proof of principle, the response selectivity of the genosensor was evaluated using PCR amplicons derived from lolB gene of Vibrio cholerae. Factors affecting immobilization, hybridization, and nonspecific binding were optimized to maximize sensitivity and reduce assay time. On the basis of the background amperometry signals obtained from nonspecific organisms and positive signals obtained from known V. cholerae, a threshold point of 4.20 microA signal was determined as positive. Under the optimum conditions, the limit of detection (LOD) of the assay was 10 CFU/mL of V. cholerae. The overall precision of this assay was good, with the coefficient of variation (CV) being 3.7% using SPCE and intermittent pulse amperometry (IPA) as an electrochemical technique. The assay is sensitive, safe, and cost-effective when compared to conventional agarose gel electrophoresis, real-time PCR, and other enzyme-linked assays for the detection of PCR amplicons. Furthermore, the use of a hand-held portable reader makes it suitable for use in the field.
    Matched MeSH terms: DNA, Bacterial/analysis*; DNA, Bacterial/genetics
  13. Polydimethylsiloxane-Paper Hybrid Lateral Flow Assay for Highly Sensitive Point-of-Care Nucleic Acid Testing
    Choi JR, Liu Z, Hu J, Tang R, Gong Y, Feng S, et al.
    Anal Chem, 2016 06 21;88(12):6254-64.
    PMID: 27012657 DOI: 10.1021/acs.analchem.6b00195
    In nucleic acid testing (NAT), gold nanoparticle (AuNP)-based lateral flow assays (LFAs) have received significant attention due to their cost-effectiveness, rapidity, and the ability to produce a simple colorimetric readout. However, the poor sensitivity of AuNP-based LFAs limits its widespread applications. Even though various efforts have been made to improve the assay sensitivity, most methods are inappropriate for integration into LFA for sample-to-answer NAT at the point-of-care (POC), usually due to the complicated fabrication processes or incompatible chemicals used. To address this, we propose a novel strategy of integrating a simple fluidic control strategy into LFA. The strategy involves incorporating a piece of paper-based shunt and a polydimethylsiloxane (PDMS) barrier to the strip to achieve optimum fluidic delays for LFA signal enhancement, resulting in 10-fold signal enhancement over unmodified LFA. The phenomena of fluidic delay were also evaluated by mathematical simulation, through which we found the movement of fluid throughout the shunt and the tortuosity effects in the presence of PDMS barrier, which significantly affect the detection sensitivity. To demonstrate the potential of integrating this strategy into a LFA with sample-in-answer-out capability, we further applied this strategy into our prototype sample-to-answer LFA to sensitively detect the Hepatitis B virus (HBV) in clinical blood samples. The proposed strategy offers great potential for highly sensitive detection of various targets for wide application in the near future.
    Matched MeSH terms: DNA, Viral/blood
  14. Graphite-Based Nanocomposite Electrochemical Sensor for Multiplex Detection of Adenine, Guanine, Thymine, and Cytosine: A Biomedical Prospect for Studying DNA Damage
    Ng KL, Khor SM
    Anal Chem, 2017 09 19;89(18):10004-10012.
    PMID: 28845664 DOI: 10.1021/acs.analchem.7b02432
    Guanine (G), adenine (A), thymine (T), and cytosine (C) are the four basic constituents of DNA. Studies on DNA composition have focused especially on DNA damage and genotoxicity. However, the development of a rapid, simple, and multiplex method for the simultaneous measurement of the four DNA bases remains a challenge. In this study, we describe a graphite-based nanocomposite electrode (Au-rGO/MWCNT/graphite) that uses a simple electro-co-deposition approach. We successfully applied the developed sensor for multiplex detection of G, A, T, and C, using square-wave voltammetry. The sensor was tested using real animal and plant DNA samples in which the hydrolysis of T and C could be achieved with 8 mol L-1 of acid. The electrochemical sensor exhibited excellent sensitivity (G = 178.8 nA/μg mL-1, A = 92.9 nA/μg mL-1, T = 1.4 nA/μg mL-1, and C = 15.1 9 nA/μg mL-1), low limit of detection (G, A = 0.5 μg mL-1; T, C = 1.0 μg mL-1), and high selectivity in the presence of common interfering factors from biological matrixes. The reliability of the established method was assessed by method validation and comparison with the ultraperformance liquid chromatography technique, and a correlation of 103.7% was achieved.
    Matched MeSH terms: DNA Damage
  15. On-Demand Ligand-Base DNA Sensor with Electrochemical Impedance Spectroscopy
    Han H, Sabani NB, Nobusawa K, Takei F, Nakatani K, Yamashita I
    Anal Chem, 2023 Jul 04;95(26):9729-9733.
    PMID: 37341999 DOI: 10.1021/acs.analchem.3c01126
    We have developed a DNA sensor that can be finalized to detect a specific target on demand. The electrode surface was modified with 2,7-diamino-1,8-naphthyridine (DANP), a small molecule with nanomolar affinity for the cytosine bulge structure. The electrode was immersed in a solution of synthetic probe-DNA that had a cytosine bulge structure at one end and a complementary sequence to the target DNA at the other end. The strong binding between the cytosine bulge and DANP anchored the probe DNAs to the electrode surface, and the electrode became ready for target DNA sensing. The complementary sequence portion of the probe DNA can be changed as requested, allowing for the detection of a wide variety of targets. Electrochemical impedance spectroscopy (EIS) with the modified electrode detected target DNAs with a high sensitivity. The charge transfer resistance (Rct) extracted from EIS showed a logarithmic relationship with the concentration of target DNA. The limit of detection (LoD) was less than 0.01 μM. By this method, highly sensitive DNA sensors for various target sequences could be easily produced.
    Matched MeSH terms: DNA/chemistry; DNA Probes
  16. Distribution of prosaposin mRNA in the central nervous system of the pigeon (Columba livia)
    Islam MR, Abdullah JM, Atoji Y
    Anat Histol Embryol, 2013 Aug;42(4):257-65.
    PMID: 22994540 DOI: 10.1111/ahe.12009
    Bioassay and immunohistochemical studies have detected the presence of prosaposin in the central nervous system (CNS) of mammals. Here, first time, we have determined the partial cDNA sequence of pigeon prosaposin and mapped the distribution of its mRNA in the pigeon CNS. The predicted amino acid sequence of pigeon prosaposin showed 93 and 60% identity to chicken and human prosaposin, respectively. In situ hybridization, autoradiograms showed that the prosaposin mRNA expression was found in the olfactory bulb, prepiriform cortex, Wulst, mesopallium, nidopallium, hippocampal formation, thalamus, tuberis nucleus, pre-tectal nucleus, nucleus mesencephalicus lateralis, pars dorsalis, nucleus isthmi, pars parvocellularis and magnocellularis, Edinger-Westphal nucleus, optic tectum, cerebellar cortex and nuclei, vestibular nuclei and gray matter of the spinal cord. These results suggest that the cDNA sequence of pigeon prosaposin is comparable to other vertebrates, and the general distribution pattern of prosaposin mRNA resembles those are found in mammals.
    Matched MeSH terms: DNA, Complementary
  17. Comparative analysis of tests used to assess sperm chromatin integrity and DNA fragmentation
    Dutta S, Henkel R, Agarwal A
    Andrologia, 2021 Mar;53(2):e13718.
    PMID: 32628294 DOI: 10.1111/and.13718
    Male infertility has a complex etiology, and many times, the cause is unknown. While routine semen analysis provides an overview of basic semen parameters, such as sperm concentration, motility, viability and morphology, a significant overlap of these parameters has been reported in fertile and infertile men. Moreover, conventional semen parameters do not reveal the cellular or molecular mechanisms of sperm dysfunctions leading to infertility. Therefore, sperm functional parameters, including sperm chromatin integrity, are evaluated to provide information on subtle sperm defects that are not routinely identified. Incomplete or defective sperm chromatin condensation increases the susceptibility of the sperm DNA to oxidative damage or other factors. To evaluate sperm chromatin integrity, different methods with varying degrees of diagnostic and prognostic capabilities are available. Among these assays, SCSA, TUNEL and SCD assays are most commonly used. While these assays rather evaluate the DNA directly for damages, the aniline blue and chromomycin A3 stains test for the quality of chromatin condensation. Thus, this review discusses and compares different methods used to evaluate sperm chromatin integrity and condensation, and their inclusion in the routine evaluation of the male infertility.
    Matched MeSH terms: DNA Fragmentation
  18. Afterword: An update on clinical utility and diagnostic value of various andrological techniques
    Durairajanayagam D, Agarwal A, Baskaran S, Vij S
    Andrologia, 2021 Mar;53(2):e13819.
    PMID: 33620116 DOI: 10.1111/and.13819
    Matched MeSH terms: DNA Damage
  19. Causes and consequences of sperm mitochondrial dysfunction
    Durairajanayagam D, Singh D, Agarwal A, Henkel R
    Andrologia, 2021 Feb;53(1):e13666.
    PMID: 32510691 DOI: 10.1111/and.13666
    Mitochondria have multiple functions, including synthesis of adenine triphosphate, production of reactive oxygen species, calcium signalling, thermogenesis and apoptosis. Mitochondria have a significant contribution in regulating the various physiological aspects of reproductive function, from spermatogenesis up to fertilisation. Mitochondrial functionality and intact mitochondrial membrane potential are a pre-requisite for sperm motility, hyperactivation, capacitation, acrosin activity, acrosome reaction and DNA integrity. Optimal mitochondrial activity is therefore crucial for human sperm function and semen quality. However, the precise role of mitochondria in spermatozoa remains to be fully explored. Defects in sperm mitochondrial function severely impair the maintenance of energy production required for sperm motility and may be an underlying cause of asthenozoospermia. Sperm mtDNA is susceptible to oxidative damage and mutations that could compromise sperm function leading to infertility. Males with abnormal semen parameters have increased mtDNA copy number and reduced mtDNA integrity. This review discusses the role of mitochondria in sperm function, along with the causes and impact of its dysfunction on male fertility. Greater understanding of sperm mitochondrial function and its correlation with sperm quality could provide further insights into their contribution in the assessment of the infertile male.
    Matched MeSH terms: DNA, Mitochondrial/genetics; DNA, Mitochondrial/metabolism
  20. Fulltext Down-regulation of steroidogenesis-related genes and its accompanying fertility decline in streptozotocin-induced diabetic male rats: ameliorative effect of metformin
    Nna VU, Bakar ABA, Ahmad A, Mohamed M
    Andrology, 2019 01;7(1):110-123.
    PMID: 30515996 DOI: 10.1111/andr.12567
    BACKGROUND: Metformin has long been used for glycemic control in diabetic state. Recently, other benefits of metformin beyond blood glucose regulation have emerged.

    OBJECTIVES: To investigate the effect of metformin on the expression of testicular steroidogenesis-related genes, spermatogenesis, and fertility of male diabetic rats.

    MATERIALS AND METHODS: Eighteen adult male Sprague Dawley rats were divided into three groups, namely normal control (NC), diabetic control (DC), and metformin-treated (300 mg/kg body weight/day) diabetic rats (D+Met). Diabetes was induced using a single intraperitoneal injection of streptozotocin (60 mg/kg b.w.), followed by oral treatment with metformin for four weeks.

    RESULTS: Diabetes decreased serum and intratesticular testosterone levels and increased serum but not intratesticular levels of luteinizing hormone. Sperm count, motility, viability, and normal morphology were decreased, while sperm nuclear DNA fragmentation was increased in DC group, relative to NC group. Testicular mRNA levels of androgen receptor, luteinizing hormone receptor, cytochrome P450 enzyme (CYP11A1), steroidogenic acute regulatory (StAR) protein, 3β-hydroxysteroid dehydrogenase (HSD), and 17β-HSD, as well as the level of StAR protein and activities of CYP11A1, 3β-HSD, and 17β-HSD, were decreased in DC group. Similarly, decreased activities of epididymal antioxidant enzymes and increased lipid peroxidation were observed in DC group. Consequently, decreased litter size, fetal weight, mating and fertility indices, and increased pre- and post-implantation losses were recorded in DC group. Following intervention with metformin, we observed increases in serum and intratesticular testosterone levels, Leydig cell count, improved sperm parameters, and decreased sperm nuclear DNA fragmentation. Furthermore, mRNA levels and activities of steroidogenesis-related enzymes were increased, with improved fertility outcome.

    DISCUSSION AND CONCLUSION: Diabetes mellitus is associated with dysregulation of steroidogenesis, abnormal spermatogenesis, and fertility decline. Controlling hyperglycemia is therefore crucial in preserving male reproductive function. Metformin not only regulates blood glucose level, but also preserves male fertility in diabetic state.

    Matched MeSH terms: DNA Fragmentation
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