A Low-Cost Micro-Volume Nephelometric System for Quantitative Immunoagglutination Assays.

Immunoassays have been widely used in scientific research and clinical diagnosis due to their versatile detection capability and high specificity. Immunoagglutination assays are kinds of immunoassay, which can simply and rapidly measure the concentration of analytes. In this work, we developed a low-cost micro-volume nephelometric system for quantitative immunoagglutination assays. We used off-the-shelf components to build the system, and the total cost of key components is only about 20 US dollars. The total detection volume in our system was as low as 3 µL, which could significantly reduce the reagent cost and required sample volume. We further evaluated the system performance via the immunoagglutination assay to measure the concentration of C-reactive protein, a plasma protein with levels rising in response to inflammation. The results demonstrated that our system could measure the concentration of analytes with relatively high sensitivity and precision within four minutes, and has high potential to be applied for clinical diagnostic tests.

Quantitative measurement of urinary proteins in 2018: advantages, disadvantages, limits.

Today, there is no reference method for the measurement of urinary proteins. The difficulties are that urine is a very complex biological fluid, and that there are a high intra-and inter-individual variability in the protein excretion rate. Progress has been made during the last thirty years, but high analytical variability persists among the colorimetric or turbidimetric methods used for urinary proteins measurement.

Rapid identification of antibiotic resistance using droplet microfluidics.

Culturing bacteria and monitoring bacterial cell growth is a critical issue when dealing with patients who present with bacterial infections. One of the main challenges that arises is the time taken to identify the particular strain of bacteria and consequently, decide the correct treatment. In the majority of cases, broad spectrum antibiotics are used to target infections when a narrow spectrum drug would be more appropriate. The efficient monitoring of bacterial growth and potential antibiotic resistance is necessary to identify the best treatment options for patients. Minturising the reactions into microfluidic droplets offers a novel method to rapidy analyze bacteria. Microfluidics facilitates low volume reactions that provide a unique system where each droplet reaction acts as an individual bioreactor. Here, we designed and built a novel platform that allowed us to create and monitor E.coli microfluidic droplet cultures. Optical capacity was built in and measurements of bacterial cultures were captured facilitating the continuous monitoring of individual reactions. The capacity of the instrument was demonstrated by the application of treatments to both bacteria and drug resistant strains of bacteria. We were able to detect responses within one hour in the droplet cultures, demonstrating the capacity of this workflow to the culture and rapid characterization of bacterial strains.

An affordable open-source turbidimeter.

Turbidity is an internationally recognized criterion for assessing drinking water quality, because the colloidal particles in turbid water may harbor pathogens, chemically reduce oxidizing disinfectants, and hinder attempts to disinfect water with ultraviolet radiation. A turbidimeter is an electronic/optical instrument that assesses turbidity by measuring the scattering of light passing through a water sample containing such colloidal particles. Commercial turbidimeters cost hundreds or thousands of dollars, putting them beyond the reach of low-resource communities around the world. An affordable open-source turbidimeter based on a single light-to-frequency sensor was designed and constructed, and evaluated against a portable commercial turbidimeter. The final product, which builds on extensive published research, is intended to catalyze further developments in affordable water and sanitation monitoring.

The usefulness of the prognostic inflammatory and nutritional index (PINI) in a haemodialysis population.

BACKGROUND AND AIM: Protein-Energy Wasting and inflammation are the principal risk factors of haemodialysis complications. We evaluated the reliability of a simple and non expensive test, the Prognostic Inflammatory and Nutritional Index (PINI), for regular screening of maintenance haemodialysis (MHD) patients in order to detect early onset of inflammation and malnutrition. METHODS AND RESULTS: 121 adult patients on maintenance dialysis were followed up for 32 months and screened every 6 months for PINI, calculated as alpha1-Acid Glycoprotein (alpha1-AG)xC-Reactive Protein (CRP)/AlbuminxTransthyretin. PINI score < or =1 was considered normal. Patients were stratified according to their PINI score: 86 patients (71.66%) had a normal score, whereas 35 (28.33%) had PINI > or = 1. The latter also had higher CRP levels, despite no clinical evidence of inflammation at the time of enrolment. Survival in patients with normal PINI was similar to patients with normal CRP, while in patients with abnormal PINI it was significantly lower than in patients with low serum albumin (p<0.05) or elevated CRP (p<0.05). After follow-up, all surviving MHD patients with PINI > or = 1 had at least one cardiovascular event vs 2.5% of patients with PINI > or = 1. CONCLUSION: The assessment of PINI can reliably identify MHD patients at higher risk of mortality and morbidity even in the absence of overt Malnutrition-Inflammation Complex Syndrome (MICS). This simple test appears to be more sensitive and specific of the single components, and not expensive, so that it could be routinely used to identify patients with sub-clinical inflammation and/or malnutrition.

Cost-effective diffuse reflectance spectroscopy device for quantifying tissue absorption and scattering in vivo.

A hybrid optical device that uses a multimode fiber coupled to a tunable light source for illumination and a 2.4-mm photodiode for detection in contact with the tissue surface is developed as a first step toward our goal of developing a cost-effective, miniature spectral imaging device to map tissue optical properties in vivo. This device coupled with an inverse Monte Carlo model of reflectance is demonstrated to accurately quantify tissue absorption and scattering in tissue-like turbid synthetic phantoms with a wide range of optical properties. The overall errors for quantifying the absorption and scattering coefficients are 6.0+/-5.6 and 6.1+/-4.7%, respectively. Compared with fiber-based detection, having the detector right at the tissue surface can significantly improve light collection efficiency, thus reducing the requirement for sophisticated detectors with high sensitivity, and this design can be easily expanded into a quantitative spectral imaging system for mapping tissue optical properties in vivo.

Screening for microalbuminuria by use of microproteinuria.

We aimed to develop a reliable, low cost method to assess the early stages of renal impairment in diabetes, for use in high-risk populations in countries with limited resources. We evaluated a trichloroacetic acid (TCA) turbidimetric method for microproteinuria screening in patients with diabetes. The method was compared with an immunoturbidimetric procedure for the detection of microalbumuniuria. Both methods performed within limits of allowable uncertainty based on inter- and intra-individual variation. A urinary albumin/creatinine ratio of 3.0 g/mol, assumed as diagnostic of microalbuminuria, was found to correlate with a cut-off value of 24 mg/L for microproteinuria. The clinical sensitivity and specificity of the TCA method determined against this ratio were 86% and 90% respectively. The reliability and practicability of the TCA method renders it suitable for the detection of early stage renal damage, with emphasis on screening high-risk populations in countries with limited resources.

Development of immunoturbidimetric assays for fourteen human serum proteins on the Hitachi 912.

Many laboratories rely on dedicated nephelometers or turbidimeters and commercial reagent kits for the evaluation of serum proteins. However, with growing emphasis on cost containment, laboratories are forced to seek additional operational efficiencies by capitalizing on the use of existing analyzers whenever possible. In the present paper we describe the development of immunoturbidimetric assays for routine analysis of 14 human serum proteins (alpha1-antitrypsin, alpha2-macroglobulin, albumin, apolipoproteins Al and B, complement components 3 and 4, haptoglobin, immunoglobulins A, G, and M, orosomucoid, prealbumin, and transferrin) on the Hitachi 912, a general chemistry analyzer. With this system, we obtained excellent precision at levels corresponding to low, normal, and high physiologic concentrations of each protein (within-run imprecision CVs < or = 3.4%, total imprecision CVs < or = 4.1%). Linearity for each method was within 5% of the expected value throughout the calibration range, and method comparisons with either the Roche turbidimetric or Dade Behring nephelometric assays were in good agreement (r >0.97). We observed no significant interference from bilirubin (up to 718 micromol/l), hemoglobin (up to 8 g/l), triglyceride (up to 14.7 mmol/l) or rheumatoid factor (up to 4,140 IU/ml). Calibration for the 14 protein assays was stable for at least 7 days and onboard refrigerated reagents were stable for at least 3 months. The instrument's automated sample re-run feature minimized sample handling and helped to conserve specimens. In conclusion, the newly developed assays on the Hitachi 912 offer high throughput (>250 tests per hour) without the associated cost of a dedicated instrument for protein assays.

Rapid monitoring of recombinant protein products: a comparison of current technologies.

Specific measurement of recombinant protein titer in a complex environment during industrial bioprocessing has traditionally relied on labor-intensive and time-consuming immunoassays. In recent years, however, developments in analytical technology have resulted in improved methods for protein product monitoring during bioprocessing. The choice of product-monitoring technology for a particular bioprocess will depend on a variety of assay factors and instrument-specific factors. In this article, we have compiled an overview of the advantages and disadvantages of the most commonly used technologies used: electrochemiluminescence, optical biosensors, rapid chromatography and nephelometry. The advantages of each technology for measuring both small and large recombinant therapeutic proteins are compared with a conventional enzyme-linked immunosorbent assay (ELISA) technique.

Changes in plasma cystatin C after renal transplantation and acute rejection in adults.

BACKGROUND: Cystatin C has recently been proposed as an alternative marker of glomerular filtration rate. The diagnostic value of plasma cystatin C for the longitudinal assessment of kidney function after renal transplantation, however, has not been addressed. METHODS: Renal function was evaluated in 30 adults receiving renal transplants (46 +/- 9 years, mean +/- SD) and in 56 healthy controls (38 +/- 10 years) using cystatin C. Plasma cystatin C was determined daily starting the day of surgery and for 3 weeks after surgery by an immunonephelometric assay. RESULTS: Plasma concentration significantly decreased during the first week (-44% vs -29% for creatinine). Plasma cystatin C correlated with plasma creatinine (r = 0.741; P <0.0001) and the reciprocal of the creatinine clearance estimated by the Cockcroft-Gault formula (r = 0.882; P <0.001). In all three cases of acute renal impairment, the increase in plasma cystatin C values was more prominent than that of creatinine. CONCLUSIONS: Plasma cystatin C is an alternative and accurate marker of allograft function in adult transplant patients. Increased sensitivity compared with creatinine for the detection of acute reduction in glomerular filtration rate allows in some cases a more rapid diagnosis of acute rejection or treatment nephrotoxicity.

Monitoring of monoclonal gammopathies: rational use of densitometry and rate nephelometry.

The follow-up of patients with monoclonal gammopathies at our institution includes serial serum protein electrophoresis (SPE) with densitometry and IgG, IgA, and IgM quantitative immunoglobulin (QIG) determinations by rate nephelometry. This retrospective audit compares monoclonal protein concentration as estimated by SPE versus QIG in 456 serial serum specimens from 105 patients to determine whether any of the tests provide redundant information. A comparison of the methods demonstrated good correlation between SPE (x-axis) and QIG (y-axis) quantitation for each immunoglobulin class: IgG had a slope of 1.45 and an intercept of 1.60 (Sy/x = 7.46, r = 0.96, n = 250); IgA had a slope of 1.30 and an intercept of -1.37 (Sy/x = 6.85, r = 0.96, n = 78); and IgM had a slope of 1.95 and an intercept of 2.06 (Sy/x = 5.16, r = 0.98, n = 128). The data for individual patients showed similar good correlations. Exceptions included IgA peaks "buried" in the beta region of the SPE (resulting in invalid SPE estimates of monoclonal protein concentration), and IgG peaks of less than 10 g/L (when background polyclonal IgG immunoglobulin skews the QIG estimate of monoclonal protein concentration). An algorithm is proposed whereby monoclonal protein concentration is measured by the specific QIG (i.e., IgG, IgA, or IgM) determination for the routine monitoring of patients, except for those with IgG peaks of less than 10 g/L that are followed by SPE.

Apoprotein assays: radial immunodiffusion versus an immunoturbidometric technique.

The measurement of apolipoprotein using radial immunodiffusion (RID) and Cobas-Bio immunoturbidometric assays is compared. Samples were obtained from 100 patients in a fiber-diet study. RID assays were performed using plates, standards, and diluent from Tago Inc. The immunonephelometric assays were performed on a Cobas-Bio with DENS program centrifugal analyzer. The correlation coefficient for the apolipoprotein Al was 0.90 and for apolipoprotein B was 0.91. The bias of the RID was to report higher values. The Cobas-Bio system affords the clinical laboratory with a reliable and cost-effective means for assessing these apolipoproteins.

Screening for slight albuminuria: a comparison of selected commercially available methods.

Slight albuminuria predicts clinically significant nephropathy in patients with insulin-dependent diabetes mellitus (IDDM) and early death in patients with non-insulin-dependent diabetes mellitus (NIDDM). This study compares four commercially available methods for measuring low concentrations of urinary albumin. We tested random spot urine specimens from 50 nondiabetic volunteers and 100 diabetic patients. This specimen was chosen to simplify collection in an outpatient setting. Two screening methods were evaluated for their ability to detect urinary albumin in the range of 15 to 200 mg/L. Sensitivity and specificity were 100% and 54.7%, respectively, for the Ames Micro-Bumintest; and 95.5% and 91.5%, respectively, for the Sclavo Albumin Screen. The high number of false-positive results made the Micro-Bumintest unacceptable. The Albumin Screen yielded fewer false-positive results, but also produced some false-negatives. Two quantitative methods, a radioimmunoassay (RIA) and a turbidimetric assay (the SPQ Microalbumin), yielded results that agreed well with each other.