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The rising prevalence of Chronic Kidney Disease (CKD) has necessitated efforts towards the development of cost-effective and accurate biosensors for serum creatinine, which is a potent biomarker reflecting kidney function. This work presents a novel and cost-effective technique to estimate serum creatinine without any sample preprocessing. The technique involves the conversion of creatinine by a monoenzymatic pathway to 1-methylhydantoin. The concentration of 1-methylhydantoin is then quantified by utilizing its innate ability to form a complex with transition metals such as cobalt. The complex formation has been validated using optical spectroscopy and the transmittance at 290 nm wavelength is used to identify the optimum concentration of cobalt chloride in sensing chemistry. This chemical assay is shown to be robust against interference from serum albumin, the abundant plasma protein that can potentially influence the sensor response. The electrochemical biosensor developed using screen-printed electrodes thus provides highly selective creatinine estimation over the range of 0.2-4 mg/dL in a sample volume of 300 μL with no preprocessing and hence can be easily translated into a viable point-of-care (POC) device.

Diabetes leads to chronic microvascular complications for the heart, kidney, and eyes due to uncontrolled glycemic fluctuations. Self-monitoring blood glucose meters can only provide a snapshot of glucose level and are incapable of capturing the granular glucose fluctuations over the 24 h in day. The clinical research has indicated that random blood glucose fluctuations can lead to organ damage. In pursuit of better glucose management, Continuous Glucose Monitoring (CGM) is emerging as a popular alternative owing to its ability to detect instantaneous changes in glucose levels and to alert the users of impending hypo- or hyper-glycemic events. In the last decade, several CGM devices have been launched in the market based on different glucose sensing chemistries and techniques. More research is still needed to come up with novel bio sensing concepts to make CGM low cost and highly accurate. Here, we elaborate the CGM techniques such as electrochemical, optical, reverse iontophoresis, microdialysis, and impedance spectroscopy. We emphasize on the widely used electrochemical CGMs with a focus on sensor design and bio-compatibility. We also provide an outlook for the future technologies, highlighting the need for innovative materials, possibility of integrating with the Internet of Things (IoT) for real-time e-health monitoring.

Aim To understand the mechanism of glycation of albumin and effects on cysteinylation and methionine oxidation. Methods The in vitro glycation of HSA and BSA was studied with varying concentrations of glucose. Clinical blood samples of diabetic subjects with varying HbA1c values, were analyzed to assess in vivo glycation. All samples and their tryptic digests were analyzed using liquid chromatography/mass spectrometry. Glycation sites were mapped on to the three-dimensional structure of the HSA and BSA. Results A total thirty-one sites for glycation and eight sites of Ne-carboxymethyl-lysine (CML) modification were identified on albumin. The site selectivity of glycation was correlated with the environment of the reactive residue in the three-dimensional structure. Conclusions The maximum percentage glycation under extreme conditions was in the range of ~55 to 88% in four weeks. Two major glycation sites K-233 and K-525 were identified, which together accounted for 40–50% of total glycation. A correlation was observed between glycation and oxidation of methionine residues in samples glycated in vitro. The role of spatially proximate residues in facilitating the glycation process is evident. The tri- and tetra-glycated isoforms of albumin can serve as biomarkers for the severe uncontrolled diabetic state.

Creatinine is a reliable indicator of renal function and degradation of muscular metabolism. Current analytical techniques for its measurement are limited by their cost and requirement of sophisticated instruments. In this work, we report a highly sensitive amperometric biosensor for creatinine by utilizing the one-step selective conversion of creatinine by creatinine deiminase. The novelty of the proposed sensor relies on the measurement of N-methylhydantoin produced in the reaction. The sensing chemistry comprises of creatinine deiminase as the receptor for creatinine, cobalt chloride as the electrochemically active recognition element, and methylene blue as the redox mediator. The sensing chemistry is immobilized on the glassy carbon electrode surface by physisorption. We have been able to provide a standalone device that reliably quantifies creatinine in serum and even whole blood, without any sample pre-processing. It is possible to measure creatinine in the clinically relevant range from 0.8 to 4 mg dL−1 with this approach.

Introduction Early detection of diabetes mellitus (DM) and diabetic kidney disease (DKD) is important for preventing end-stage renal failure and reducing cardiovascular complications. Availability of a validated point-of-care (PoC) device that can measure various DKD markers would be useful in this respect, especially in resource-poor parts of the world. Methods We validated a novel nanotechnology-based multianalyte PoC device (minimally invasive and does not require trained medical personnel) against laboratory gold standard tests for the detection of 5 biomarkers related to management of DM and DKD. The prospective study was funded by an International Society of Nephrology American Nephrologists of Indian Origin grant in 2 phases: (i) proof of concept: random samples were tested for the analytes with the PoC device and correlated with the laboratory gold standard; and (ii) clinical validation in a well-characterized cohort of patients. A nonenzymatic- and nonantibody-based electrochemical PoC device for quantitative measurement of markers—glycosylated hemoglobin (HbA1c), hemoglobin, serum albumin, microalbuminuria, urine creatinine, and albumin-to-creatinine ratio−was developed and used in this study. The disposable strips were interfaced with a multipotentiostat hand-held PoC device (3.7-V rechargeable lithium battery, 5-inch touch screen, Bluetooth enabled) working in amperometry mode, which provided the results in

Serum creatinine is an established biochemical marker for monitoring of renal function. Analytical methods to detect creatinine concentration have been investigated as early as 1886. However, majority of these methods involve sophisticated instrumentation which limits its accessibility to economically challenged patients. In this regard, point of care biosensors provide a low-cost solution coupled with simple instrumentation. There has been growing interest in development of point-of-care technology (POCT) devices for creatinine estimation. However, majority of these POCT devices are enzymatic assays, which add to the complexity and the cost. In this work, we aim to develop non-enzymatic electrochemical creatinine biosensor which simplifies the process of routine clinical analysis, and thereby making it accessible due to its low-cost.

A non-enzymatic electrochemical technique for creatinine sensing is presented, exploiting iron binding property of creatinine. Disposable carbon printed electrodes layered with FeCl3 coated cotton fiber membranes are used to sense creatinine from 10 to 245 mg/dl, on clinical urine samples. The energy-dispersive X-ray spectroscopy analysis confirms the presence of Fe(III) dry chemistry on cotton membrane. Creatinine binding with Fe(III) is verified with UV analysis, with a corresponding decrease in Fe(III) reduction current in cyclic voltammetry. The disposable test strips are interfaced with multi-potentiostat point of care (POC) hand-held device, working in amperometry mode. The results obtained on POC biosensors demonstrate good correlation ( $R^{2}=0.91$ ) with Jaffe method laboratory gold standard. The intra-assay variability is less than 7.1%. The statistical bias as revealed from the Bland–Altman analysis indicates that the POC results are within 95% confidence interval. This POC device does not require any sample preparation step and provides sample to result in less than a minute. FeCl3 sensing chemistry is robust against urine albumin interference, which is especially significant for accurate estimation of albumin to creatinine ratio. The non-enzymatic nature of disposable test strips results in highly stable and robust operation of the POC device over a large range of temperature variations.

Leptospirosis is a recurring global disease of severe illness involving pulmonary and renal involvement in cattle and humans that needs attention to cure. The major challenge in treating leptospirosis disease is the diagnosis of the disease. The culturing of an organism is a gold standard for confirmation of the disease. The growth and optimistic identification of an organism require at least 8 to 14 days because of its slow growth characteristics. We have investigated various media conditions that are prepared based on the wealth of information obtained from 'omic' studies and report a sustainable Leptospira growth medium comprising of serum equivalent elements and nutrients for pyrimidine biosynthesis, which allows a visible growth of organism within 2 days.

Direct Electron Transfer biosensors, facilitating direct communication between the biomolecule of interest and electrode surface, are preferable compared to enzymatic and mediator based sensors. Although hemoglobin (Hb) contains four redox active iron centres, direct detection is not possible due to inaccessibility of iron centres and formation of dimers, blocking electron transfer. Through the coordination of iron with aza-heterocyclic receptors - pyridine and imidazole - we report a cost effective, highly sensitive and simple electrochemical Hb sensor using cyclic voltammetry and chronoamperometry. The receptor can be either in the form of liquid micro-droplet mixed with blood or dry chemistry embedded in paper membrane on top of screen printed carbon electrodes. We demonstrate excellent linearity and robustness against interference using clinical samples. A truly point of care technology is demonstrated by integrating disposable test strips with handheld reader, enabling finger prick to result in less than a minute.

Using a combination of avidin-biotin microELISA and solid phase radioimmunoassay, we examined sera from 23 patients with systemic lupus erythematosus (SLE), two patients with established sensitivity to ingested shrimp, and 15 healthy normal subjects. In addition to IgG antibodies, varying amounts of IgE antibodies specific for native DNA (nDNA), denatured or single-stranded DNA (dnDNA), RNA, and tRNA were demonstrable in the sera of SLE patients, but not in the sera of normal subjects. A comparison of the specificity of nucleic acid-specific IgE antibodies present in the sera of shrimp-sensitive patients with those present in the sera of seven SLE patients revealed that the IgE antibodies in the sera of shrimp-sensitive patients specifically recognized shrimp tRNA but not yeast tRNA, calf thymus RNA, or calf thymus DNA, while those present in the sera of patients with SLE recognized all these nucleic acid antigens. The IgE antibodies directed against nDNA, dnDNA, RNA, and tRNA may mediate mast cell and basophil degranulation and thus contribute both to immediate-type hypersensitivity phenomena including hives seen in patients with SLE and to the localization of IgE-nucleic acid complexes in target tissues.

The affinity of phosvitin with serine hydroxymethyl transferase (SHMT), an acidic multi-subunit protein, was evaluated by measurements of enzyme activity, sedimentation velocity, steady-state fluorescence, circular dichroism and kinetic thermal stability. While the presence of phosvitin had no effect on the SHMT activity, the sedimentation coefficient of SHMT increased from 8.7 S to 12.5 S suggesting the formation of a complex at a SHMT:phosvitin molar ratio of 2:1. Based on steady-state fluorescence quenching measurements an association constant of 2.4 +/- 0.2 x 10(5) M-1 at 25 degrees C was obtained for the interaction of phosvitin with SHMT. The temperature dependency of the association constant in the range 15-35 degrees C suggests the involvement of ionic forces in the interaction. The thermal inactivation of SHMT followed first order kinetics. In the presence of phosvitin the rate constant decreased and half time increased. The circular dichroism measurements suggest that phosvitin interaction does not involve pyridoxal phosphate binding domain of the enzyme. Although minor changes in the secondary structure of the enzyme were observed, the environment around aromatic amino acids did not change significantly.

The oxidation of linoleic acid catalyzed by Fe(II) is strongly inhibited by phosvitin, while chelation with EDTA, NTA or deferoxamine produced only partial inhibition. Interestingly, the DNA degradation catalyzed by Fe(II) in the presence of H2O2 is also inhibited by phosvitin or deferoxamine. In contrast, chelation of the metal ion with EDTA or NTA enhanced the DNA degradation. The results suggest that the nature of interaction between the metal ion and the complexing agent may be an important factor in the generation of active oxygen intermediates.

The dispersion coefficient, the time delay and the time constant, as parameters associated with models of liquid phase axial dispersion in fluidized systems, have been correlated using available literature data for fluidization involving a wide range of fluid and particle properties. The results permit prediction of these parameters from only a knowledge of the bed voidage. A comparison with Chung and Wen's1 correlation demonstrates the superiority of the present correlation. On a etabli une correlation impliquant le coefficient de dispersion, le retard et la constante de temps comme parametres associes a des moddeles de dispersion axiale en phase liquide dans des systemes fluidise on a employe a cette fin les informations publiees dans la litterature sur la fluidisation, qui portent sur de larges domaines des proprietes des fluides et des particules. Les resultats obtenus permettent de prevoir ces parametres aa partir de la seule connaissance du degre de vide du lit. Une comparaison entre cette correlation et celle de Chung et Wen(1) deniontre la superiorite de la premiere.

Four methods are proposed for estimating the dominant time constant and dead time of a given process from a knowledge of its moments, s-plane, frequency or transient response data. The methods are faster and simpler and, in terms of accuracy and reliabiltiy, are comparable to the time domain least squares procedure for parameter estimation. Applications of the methods are illustrated with the aid of a dispersion model. A comparison is also made with other known techniques to demonstrate the superiority of the suggested methods.

1. Cell-free extracts of Escherichia coli K12 catalyse the synthesis of N-acetyl-l-phenylalanine from acetyl-CoA and l-phenylalanine. 2. The acetyl-CoA–l-phenylalanine α-N-acetyltransferase was purified 160-fold from cell-free extracts. 3. The enzyme has a pH optimum of 8 and catalyses the acetylation of l-phenylalanine. Other l-amino acids such as histidine and alanine are acetylated at slower rates. 4. A transacylase was also purified from E. coli extracts and its substrate specificity studied. 5. The properties of both these enzymes were compared with those of other known amino acid acetyltransferases and transacylases.

Fluid mixing in liquid fluidized beds has been characterized by a dynamic response technique using pulse inputs. Solid particles with a size range of 0.5 to 3.0 mm diameter and a density range from 1.28 to 7.02 gms/cc were fluidized with water at void fractions of 0.4 to 0.85 in a 2-inch column. A dispersed plug flow model has been applied successfully, including a frequency response comparison, and dispersion coefficients have been calculated by the moments method. The results are related to correlations previously advanced, including the use of an eddy diffusivity and of the Bodenstein number, and a more recent refined correlation allowing for density differences. On a caracterise le melange de fluides dans des lits liquides fluidises, au moyen d'une methode dynamique de reponse dans laquellc on introduit des impulsions. On a fluidise des particules solides (dont le diametre variait entre 0.5 et 3.0 millimetres et la densite etait comprise entre 1.28 et 7.02 g/c3) avec de l'eau, a des fractions de vide de 0.4 a 0.85, dans une colonne de 2 ponces. On y a applique avec succes mi modele d'ecoulement disperse avec obturation, dans lequel on coparait les reponses a la frequence, et l'on a calcule les coefficients de dispersion par la methode des moments. On apparente les resultats aux correlations deja avancees, (comprenant l'emploi de la diffusivite d'un tourbillon et du nombre de Bodenstein) ainsi qu'a une correlation subtile et plus recente, dans laquelle on tient compte des differences des densites.

The infrequency of multiple sclerosis in India may have genetic implications. We found (a) the HLA-A3 and HLA-B7 haplotypes amongst Indians to be lower than those reported in Caucasians, (b) no excess of HLA-A3 and HLA-B7 amongst our 27 multiple sclerosis patients compared to 330 controls; instead it was the reverse, (c) HLA-B12 as high as 74% in the "clinically definite" cases, against only 9% in controls, (d) a significant relative risk of MS amongst Indians with HLA-B12 haplotype. Attention is drawn to th higher incidence of MS amongst the small Parsee community and the high association of HLA-B12 in these patients.

A method of describing systems by sets of second-order differential equations is suggested. In this second-order state-space formulation the states are defined in a manner slightly different from the conventional one. It is shown that this modified procedure reduces the order of matrices, renders easier the evaluation of the characteristic equation and the system solution, and provides some additional insight into the relative stability of system states. Procedures of characterization and analysis of systems by this modified method are indicated, and an example is considered to illustrate the advantageous features of the method in contrast with the normal procedure.

gamma-Glutamyl transpeptidase (gamma-GT) activity, which catalyzes the transfer of the "gamma-glutamyl" group of gamma-glutamyl compounds to several dipeptide and amino acid acceptors, was found to be present in several mycobacteria, including M. leprae, both in cell suspensions and in cell-free sonicates. Glycyl D-amino acids were active as acceptors, particularly glycyl-D-alanine and alpha, epsilon-diaminopimelic acid, among the amino acids. Two mycobacterial isolates obtained from biopsy material of lepromatous patients also exhibited similar enzyme activity. The need for further work to delineate the possible role of gamma-GT in mycobacterial metabolism is strongly indicated.

Chemical Examination of the Roots of \(Aristolochiα\) \(Indicα\), Linn. Part I.&nbsp

An immunoscreening approach was used to isolate a strongly positive cDNA clone from an Entamoeba histolytica HK-9 cDNA expression library in the phage vector lambda ZAP-II. The 1.85-kb cDNA insert was found to be truncated and encoded the cysteine-rich, immunodominant domain of the antigenic 170-kDa subunit of the amebal galactose-N-acetylgalactosamine binding lectin. This domain was expressed as a glutathione S-transferase fusion protein in Escherichia coli. Inclusion bodies of the recombinant protein were solubilized with Sarkosyl, and the protein was enriched from the crude bacterial extract by thiol-affinity chromatography. The recombinant protein was used to develop a rapid, sensitive, and specific avidin-biotin microtiter enzyme-linked immunosorbent assay (ELISA) for invasive amebiasis. Sera from 38 individuals suffering from invasive amebiasis, 12 individuals with noninvasive amebiasis, 44 individuals with other infections, and 27 healthy subjects were screened by the recombinant antigen-based ELISA. The sensitivity and specificity of the assay were 90.4 and 94.3%, respectively, which correlated well with those of an ELISA developed with crude amebal antigen (r = 0.94; P < 0.0001), as well as with those of a commercially available serodiagnostic ELISA (r = 0.92; P < 0.0001). Thus, the bacterially expressed recombinant lectin can replace the crude amebal extract as an antigen in the serodiagnosis of invasive amebiasis by using avidin-biotin microtiter ELISA.

Die Degradationsprodukte, die sich bei der Hydrolyse der Purine, der Pyrimidine und der Nukleinsauren bilden, werden mittels runder Filterpapierscheiben chromatographisch untersucht. Adenine, Nukleinsauren und Adeninverbindungen zeigen bei einer Saurehydrolyse ausser einem Band, das den Platz des Glycin einnimmt, noch ein weiteres Band. Dieses Band ist versuchsweise als 4(5)-amino-5(4)-imidazol Carboxamidin charakterisiert worden. Die Bedeutung dieser Ergebnisse fur die quantitative Analyse der Nukleinsauren und Adeninverbindungen wird in der vorliegenden Arbeit erortert.

THE diazotizable amine which accumulated in cultures of Escherichia coli during sulphonamide bacteriostasis has been isolated and its identity with 4-amino 5-imidazole carboxamide has been established1,2. Evidence to show that this compound was not merely a product of aberrant metabolism but a normal precursor of purines has been obtained3 and its role in purine biosynthesis is well established4. Recently, it was demonstrated that the carboxamide was rapidly utilized in rat liver homogenates and preparations from actively growing tissues5,6. The present report deals with the preliminary experiments carried out to assess the role of this compound as a potential precursor of purines in plants. Resting, as well as germinating, seeds of Phaseolus radiatus (green gram) were used in this investigation.