Male Holtzman rats weighing 180–210 g (about 50 days old) were used for induction of adjuvant arthritis. Polyarthritis was induced by injection in the left hind paw of 0.1 ml of Freund’s adjuvant containing 5.0 mg ml−1 of heat inactivated Mycobacterium tuberculosis, derived from the human strain H37Rv, suspended in mineral oil (Pearson and Wood 1963 (link)). Monoarthritis was induced in the same manner but with 0.1 ml of Freund’s adjuvant containing only 1.0 mg ml−1 of M. tuberculosis (Sigma-Aldrich, St Louis, MO, USA) (Bracht et al. 2012 (link)). Rats of similar ages served as controls. After 18 days of the adjuvant injection, rats were selected for the experiments. All procedures were done in accordance with the world-wide accepted ethical guidelines for animal experimentation and previously approved by the Ethics Committee for Animal Experimentation of the University of Maringá (Protocol 094/2013-CEEA).
M tuberculosis
Manufactured by Merck Group
Sourced in United States
The M. tuberculosis lab equipment is used for the detection and identification of the Mycobacterium tuberculosis bacterium, the causative agent of tuberculosis. The equipment provides a standardized and reliable method for the analysis of clinical samples, allowing for accurate diagnosis and disease monitoring.
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5 protocols using m tuberculosis
1
Induction of Adjuvant-Induced Arthritis in Rats
2
Schistosoma japonicum Infection Model
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S. japonicum cercariae were maintained in Oncomelania hupensis snails as the intermediate host, which were purchased from Jiangsu Institute of Parasitic Disease (Wuxi, China). Each mouse was infected with 12±1 cercariae of S. japonicum by abdominal skin exposure. In each independent experiment, 12 mice were randomly chosen from the infected and normal control groups at 8 weeks post infection, and killed for further study.
For other pathogen challenge experiments, each mouse was challenged by i.p. injection of heat-inactivated M. tuberculosis (Sigma-Aldrich, St Louis MO, 4 mg kg−1). Splenocytes and peritoneal macrophages were prepared at 24 h after injection for Th1/Th2 and M1/M2 analysis.
SEA was obtained from purified and homogenized S. japonicum eggs. The protein concentration of SEA was determined using a bicinchoninic acid Protein Assay kit (Bio-Rad, Richmond, CA).
For other pathogen challenge experiments, each mouse was challenged by i.p. injection of heat-inactivated M. tuberculosis (Sigma-Aldrich, St Louis MO, 4 mg kg−1). Splenocytes and peritoneal macrophages were prepared at 24 h after injection for Th1/Th2 and M1/M2 analysis.
SEA was obtained from purified and homogenized S. japonicum eggs. The protein concentration of SEA was determined using a bicinchoninic acid Protein Assay kit (Bio-Rad, Richmond, CA).
3
Myosin-Induced Autoimmune Response in Guinea Pigs
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For guinea pigs in group A, injection of 0.01 M PBS + complete Freund adjuvant (CFA) was performed on a weekly basis. After 6 immunizations, saline was administrated 4 times.
Group B animals received an injection of myosin in 0.01 M PBS + CFA on a weekly basis. After 6 immunizations, saline was administrated 4 times.
For group C, injection of myosin in 0.01 M PBS + CFA on a weekly basis was performed. After 6 immunizations, defibrase was administrated 4 times.
Guinea pigs were immunized by subcutaneous injections of 0.25 ml purified rabbit myosin (10.8 mg/ml (Biuret), 0.75 units/mg protein, pH 6.8, Sigma-Aldrich, St. Louis, MO, USA) emulsified with an equal volume of conventional CFA containing 1 mg/ml M. tuberculosis (Sigma-Aldrich, St. Louis, MO, USA) in multiple sites of the back on a weekly basis 6 times. At the time of immunization, animals received an intraperitoneal injection of 0.3 ml of pertussis bacterium solution (2 × 109/ml pertussis bacterium, Tiantan Bio Ltd., Beijing, CN).
Group B animals received an injection of myosin in 0.01 M PBS + CFA on a weekly basis. After 6 immunizations, saline was administrated 4 times.
For group C, injection of myosin in 0.01 M PBS + CFA on a weekly basis was performed. After 6 immunizations, defibrase was administrated 4 times.
Guinea pigs were immunized by subcutaneous injections of 0.25 ml purified rabbit myosin (10.8 mg/ml (Biuret), 0.75 units/mg protein, pH 6.8, Sigma-Aldrich, St. Louis, MO, USA) emulsified with an equal volume of conventional CFA containing 1 mg/ml M. tuberculosis (Sigma-Aldrich, St. Louis, MO, USA) in multiple sites of the back on a weekly basis 6 times. At the time of immunization, animals received an intraperitoneal injection of 0.3 ml of pertussis bacterium solution (2 × 109/ml pertussis bacterium, Tiantan Bio Ltd., Beijing, CN).
4
CFA-Induced Mechanical Hypersensitivity
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Inflammation was induced by intraplantar injection of 10 μL of complete Freund’s adjuvant (CFA; 0.5 mg/mL heat-killed M. tuberculosis; Sigma, St. Louis, MO, USA) into the hind paw. Saline (pH 7.4) served as control. Mechanical sensitivity was measured using electronic von Frey filament at multiple time points (6 h to 1 week) after CFA injection.
5
Murine Model of Experimental Autoimmune Encephalomyelitis
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EAE was induced in both male and female mice between 8 and 12 weeks of age. MOG35-55 peptide (100 μg, CSBio; CS0681) was emulsified in complete Freund’s adjuvant containing Mycobacterium tuberculosis (1 mg/mL, Sigma; F5881; M. tuberculosis (BD 231141) added to final concentration of 4 mg/mL) and was injected subcutaneously (100 μL volume) at the base of the tail. Pertussis toxin (200 ng, List Biologicals; 180) was administered i.p. on the day of and 1 day after MOG immunization. Mice were scored daily by 2 blinded evaluators using the following scale: 0-no clinical disease, 1-limp tail, 2-hindlimb incoordination, 3-hindlimb weakness, 4-hindlimb paralysis, 5-moribund. As is standard practice, animals that did not develop any signs of EAE were excluded from analysis. Total incidence including excluded animals is reported in S1 Data
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