A midstream morning urine sample was collected using BD Vacutainer system (Beckton, Dickinson & Co) and transferred to the laboratory for processing. Nine mL of urine was taken from the vacutainer using a separate vacuette, which was later processed into the biobank. A small amount (10-20 µl) of urine was used for the detection of S. mansoni, using the point-of-care circulating cathodic antigen cassette (ICT Diagnostics), following the manufacturer’s instructions [64 (link)], resulting in the following classification: negative or three levels of positives (1+, 2+, or 3+). Dipstick urinalysis to detect proteins, blood, glucose, urobilinogen, bilirubin, leucocytes, ketones, nitrites, pH, and specific gravity was applied using the Roche Combur-10 test (Roche Diagnostics). Urine samples testing positive for blood were subjected to filtration test for the diagnosis of S. haematobium. Positive results were expressed as the number of S. haematobium eggs per 10 mL of urine, according to World Health Organization guidelines [65 (link)].
Combur 10 test
Manufactured by Roche
Sourced in Switzerland
The Combur 10 Test is a urine test strip manufactured by Roche. It is designed to detect the presence and concentration of various analytes in urine samples, including glucose, protein, blood, pH, and more.
Automatically generated - may contain errors
Lab products found in correlation
2 d quant kit, by GE Healthcare (3 mentions)
Urine monovette, by Sarstedt (1 mentions)
Eclipse e200mv r, by Nikon (1 mentions)
Nuclepore track etch membrane filtration products, by Cytiva (1 mentions)
Whatman, by Cytiva (1 mentions)
Urovysion assay, by Abbott (1 mentions)
Ureal, by Roche (1 mentions)
Urea nitrogen colorimetric detection kit, by Thermo Fisher Scientific (1 mentions)
Gluc3, by Roche (1 mentions)
Albt2, by Roche (1 mentions)
Crej2, by Roche (1 mentions)
Bwb 800, by Tanita (1 mentions)
Polar sports tester, by Polar Electro (1 mentions)
14 protocols using combur 10 test
1
Urine Sample Collection and Analysis for Schistosomiasis
2
Diagnosis of Parasitic Infections
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Stool samples were examined using the Baermann technique for the diagnosis of Strongyloides stercoralis [19 ] and the Kato-Katz technique [20 ] for S. mansoni and soil-transmitted helminths. Duplicate Kato-Katz thick smears were prepared from each stool sample. Within an hour after preparation, the Kato-Katz thick smears were examined under a microscope by experienced laboratory technicians for the diagnosis of hookworm eggs as the eggs deteriorate rapidly. The same thick smears were re-examined later for the diagnosis of the eggs of S. mansoni, Ascaris lumbricoides and Trichuris trichiura.
Urine samples were examined for the presence of S. haematobium eggs using a filtration method [21 ], for S. mansoni infection using a point-of-care circulating cathodic antigen (POC-CCA) cassette test (ICT Diagnostics; Cape Town, South Africa) and for microhaematuria using a Roche Combur-10 test (Roche Diagnostics; Basel, Switzerland). In accordance to the manufacturer’s instructions, POC-CCA tests were scored as either negative or positive, the latter stratified into trace, 1 +, 2 + or 3 + .
Blood samples were subjected to a rapid diagnostic test (ICT Diagnostics; Cape Town, South Africa) and thick and thin blood films were examined under a microscope for the assessment of Plasmodium infection. Haemoglobin level in blood samples was measured using Hemocue 301 system (Angelholm, Sweden).
Urine samples were examined for the presence of S. haematobium eggs using a filtration method [21 ], for S. mansoni infection using a point-of-care circulating cathodic antigen (POC-CCA) cassette test (ICT Diagnostics; Cape Town, South Africa) and for microhaematuria using a Roche Combur-10 test (Roche Diagnostics; Basel, Switzerland). In accordance to the manufacturer’s instructions, POC-CCA tests were scored as either negative or positive, the latter stratified into trace, 1 +, 2 + or 3 + .
Blood samples were subjected to a rapid diagnostic test (ICT Diagnostics; Cape Town, South Africa) and thick and thin blood films were examined under a microscope for the assessment of Plasmodium infection. Haemoglobin level in blood samples was measured using Hemocue 301 system (Angelholm, Sweden).
3
Comprehensive Urinalysis Workflow
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All the urine samples were kept refrigerated (+4 °C) and were processed on a routine basis within 2 h after collection. In particular, the urinalysis consisted of a macroscopic examination evaluating the color and turbidity. Urine specific gravity (USG) was measured using a manual refractometer (Giorgio Bormac, 41012 Modena, Italy), the chemical evaluation was carried out using a semi-quantitative dipstick test (Combur10Test, Roche Diagnostic, Mannheim, Germany). After centrifugation at 1500 g for 10 min, urine sediment at T0 was observed under both high (400x) and low microscopic fields (100x). Urine supernatants were divided into aliquots and stored in part at −20 °C for a maximum of 7 days for total protein and creatinine determination, and in part at − 80 °C for the subsequent proteomic analysis.
4
Urine Sample Preparation for Proteomics
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Midstream spot urine samples (50–100 mL)
were obtained from
participants after a standard 10 h fasting, into a sterile urine container
and transported immediately on ice to prevent microbe contamination
and proteolysis. Urine analysis was carried out to determine the presence
of urinary protein, infection, sugar, or occult blood using a urine
test strip (Combur10 Test, Roche).
The samples were then processed
and insoluble materials were removed by centrifugation at 2000g (4000 rpm) at 4 °C for 10 min, within 30 min of collection,
to prevent protein release from these artifacts. The supernatant (5
mL) was carefully removed, aliquoted, and stored at −80 °C
for long-term storage. Proteins were isolated from the urine samples,
as described previously.26 (link) The protein
pellets were solubilized in labeling buffer (7 M urea, 2 M thiourea,
30 mM Tris–HCl, and 4% 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate
(CHAPS), pH 8.5). Insoluble material was pelleted by centrifugation
(12 000g, room temperature (RT), 5 min), and
protein concentrations were determined in triplicate using the 2D-Quant
kit (GE Healthcare).
were obtained from
participants after a standard 10 h fasting, into a sterile urine container
and transported immediately on ice to prevent microbe contamination
and proteolysis. Urine analysis was carried out to determine the presence
of urinary protein, infection, sugar, or occult blood using a urine
test strip (Combur10 Test, Roche).
The samples were then processed
and insoluble materials were removed by centrifugation at 2000g (4000 rpm) at 4 °C for 10 min, within 30 min of collection,
to prevent protein release from these artifacts. The supernatant (5
mL) was carefully removed, aliquoted, and stored at −80 °C
for long-term storage. Proteins were isolated from the urine samples,
as described previously.26 (link) The protein
pellets were solubilized in labeling buffer (7 M urea, 2 M thiourea,
30 mM Tris–HCl, and 4% 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate
(CHAPS), pH 8.5). Insoluble material was pelleted by centrifugation
(12 000g, room temperature (RT), 5 min), and
protein concentrations were determined in triplicate using the 2D-Quant
kit (GE Healthcare).
5
Urine Sample Preparation for Proteomic Analysis
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After a standard 10 h fast, midstream spot urine samples (50–100 mL) were collected in a clean-catch specimen into a sterile urine container and immediately transported in ice to avoid microbial contamination and proteolysis. We used urine test strips to screen for urinary protein, urinary infection, urinary sugar, and occult blood (Combur10Test, Roche: Basel, Switzerland). Within 30 min of collection, the samples were processed, and insoluble materials were removed by centrifugation at 2000× g (4000 rpm) at 4 °C for 10 min to avoid protein release from these artifacts. For long-term storage, the supernatants were carefully removed and frozen in 2 mL aliquots at −80 °C. Proteins were extracted from urine samples as previously reported [41 (link)]. Centrifugation (12,000× g, room temperature, 5 min) was used to pellet insoluble material. In a labeling buffer, the pellets were solubilized (7 M urea, 2 M thiourea, 30 mM Tris-HCl, 4% CHAPS, pH 8.5). The 2D-Quant Kit was then used to determine the concentration of protein samples in triplicate (GE Healthcare, Chicago, IL, USA).
6
Diagnosis of Urogenital Schistosomiasis
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Urine test strip and urine filtration technique were performed on each urine sample. A urine test strip (Combur-10-Test, Roche Diagnostic, Cobas) was performed to assess the presence of haematuria. The lower detection limit of blood (erythrocyte) was 5 erythrocytes/μl. For the diagnosis of urogenital schistosomiasis, urine samples were subjected to the filtration technique and read by microscopists unware of study treatment. Urine filtration technique consisted of 10 mL of urine freshly collected injected through a chamber containing a 12μm pore-size nuclepore filter followed by microscopic examination for the detection of S. haematobium eggs as described elsewhere [24 (link),25 (link)]. The counting of eggs is done using x40 magnification scanning completely the filter in four passes and the egg count per volume of urine is recorded. Samples were considered S. haematobium positive if at least one egg was detected in at least one urine sample out of at least two urine samples provided by each subject and as negative if all two or more urine samples did not have any eggs present.
7
Urine Collection and Analysis for SCI
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Approximately 30 ml of midstream urine were collected from the controls and the study participants with SCI either using a sterile urine cup and a urine monovette (Sarstedt) or directly from a catheter (intermittent, indwelling or suprapubic). Urine was kept at 4 °C for a maximum of 5 h before being processed. A urine sample was analyzed in a certified clinical laboratory (at the Swiss Paraplegic Centre in Nottwil, Switzerland) for creatinine concentration and tested with a urine-Stix test (Combur 10 Test, Roche) with automated result acquisition (cobas u 4111). In case of a positive urine-Stix signal, a leucocyte and microbial quantification of the urine sediment was performed. If more than 90 leucocytes/ μl or an elevated microorganism count (> 1*105 /ml was detected, a subsequent bacteriological analysis was conducted to identify and characterize the infection causing bacteria. The remaining urine was centrifuged at 4 °C and 1′800 g for 10 min. The supernatant was aliquoted and stored at −80 °C until further usage. The urine sediment pellet was resuspended in 1 ml of residual urine and also frozen at −80 °C.
8
Comprehensive Joint Aspiration Analysis
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After successful aspiration, the fluid was centrifugated at 1000 rpm for 10 minprior to being tested. Then one drop of supernatant was applied to a LE test strip (Combur 10 Test, Roche Diagnostics, Mannheim, Germany). According to the four colour grades on the box (negative, ~ 10–25, ~ 75, ~ 500 leukocytes/µl), the result could be read from the color patch change on the LE strip after 2 min. Furthermore, the aspirates were tested for synovial leukocyte cell count, PMN (%), and CRP in our laboratory. Alpha-Defensin samples were sent on the same day to a collaborating laboratory (Labor Dr. Fenner and colleagues, Hamburg, Germany) where a standard enzyme-linked immunosorbent assay (ELISA) was performed. In addition, joint aspirates were applied to blood culture mediums (aerobic and anaerobic) and a microbiological swab tube with following incubation for 14 days.
9
Schistosoma haematobium Diagnosis Protocol
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Clinical assessment consisted of a questionnaire on present or previous symptoms of UGS, known bladder or kidney disease, previous praziquantel treatment, the duration of fresh water contact and pregnancy. Three urine samples, provided between 10:00 h and 14:00 h on three consecutive days, were requested. Semiquantitative urine dipstick testing as well as urine filtration (10 ml urine) for microscopy for the presence of Schistosoma haematobium were performed (Dipstick: Combur 10 Test, Roche Diagnostics Ltd, Risch, Switzerland; Millipore filter: Nuclepore Track-Etch Membrane Filtration Products, Whatman, Maidstone, UK; microscope: Eclipse E200MV R, Nikon, Tokyo, Japan). A positive result was defined as any detection of eggs in the filtered urine. Heavy intensity infection was defined as ≥50 eggs/10 ml urine. 21 In addition, 10 ml of urine was centrifuged for 5 min at 710 x g and, after discarding the supernatant, 500 μl of urine was stored at -20 • C for Schistosoma genus PCR. In the case of negative microscopy, urine real-time PCR for Schistosoma-specific DNA was performed at CERMEL, as published previously. 22
10
Urine-based Bladder Cancer Screening
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In freshly voided urine, creatinine was determined with the enzymatic test CREA plus® and leukocytes, erythrocytes and other parameters were quantified with Combur10 Test® strips (Roche Diagnostics, Mannheim, Germany). Additionally, erythrocytes and leukocytes were detected microscopically in the cell segment as previously described [22 (link)]. Urine-status data were documented for 84% of all samples. CNVs were determined with the UroVysion™ assay according to the protocol of the manufacturer (Abbott Laboratories, Abbott Park, IL). In brief, about 25 to 30 morphologically suspicious cells were evaluated in each urine sediment [18 ]. The UroVysion™ Bladder Cancer Kit is composed of three centromere-specific probes (CEP 3, CEP 7, CEP 17) to capture aneusomy of chromosomes 3, 7, and 17, and the locus-specific indicator probe (LSI) to assess CNV at 9p21. The test was considered to be positive following the manufacturer’s decision rule if ≥4 cells showed polysomy (CNV > 2) of ≥2 chromosomes (3, 7 or 17) or if ≥12 cells had no signal at 9p21 indicating loss of both alleles (CNV = 0).
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