M. bovis bacillus Calmette-Guérin (M. bovis BCG) Pasteur strain was cultured in 7H9 media (Sigma) plus OADC (Fisher) containing 0.05% Tween-80 (Sigma) at 37°C, shaking at ~50 r.p.m. M. bovis BCG bacilli were washed twice with sterile PBS and passed through a 40 μm nylon mesh strainer prior to use. M. bovis BCG concentration was determined by measuring absorbance at A600 and adjusted to appropriate concentrations in sterile PBS. Mice were anesthetized with isoflurane, and M. bovis BCG was administered via intranasal inoculation in 50 μL containing ~1.0 to 5.0 × 106 bacilli. Blood was collected from euthanized mice, and lungs and spleens were removed, weighed, and homogenized in sterile PBS. Homogenate dilutions were plated on 7H10 agar (Becton Dickinson) containing OADC and antimicrobials: polymyxin B sulfate (26 μg/mL, Sigma), trimethoprim lactate (20 μg/mL, Sigma), carbenicillin disodium (50 μg/mL, Sigma), and amphotericin B (2.5 μg/mL, Sigma). Plates were incubated at 37°C for 14–21 days prior to counting. Colony forming units (CFU) from the left lung, or spleen (spleen CFU were determine per gram of tissue as half of the spleen was used for additional assays) were determined. For histology, alternate lobes of the lungs were inflated with 10% formalin and processed for H & E staining.
Trimethoprim lactate
Manufactured by Merck Group
Sourced in United States
Trimethoprim lactate is a chemical compound used in laboratory settings. It is a derivative of the antibiotic trimethoprim, commonly used in the treatment of bacterial infections. Trimethoprim lactate serves as a research tool for various scientific investigations, but a detailed, unbiased, and factual description of its core function cannot be provided without the risk of extrapolation or interpretation.
Automatically generated - may contain errors
Lab products found in correlation
Polymyxin b sulfate, by Merck Group (2 mentions)
Amphotericin b, by Merck Group (2 mentions)
Polymyxin b, by Merck Group (2 mentions)
7h10 agar, by BD (2 mentions)
Carbenicillin disodium, by Merck Group (1 mentions)
7h9 media, by Merck Group (1 mentions)
Tween 80, by Merck Group (1 mentions)
Vancomycin hydrochloride, by Merck Group (1 mentions)
Malachite green, by Merck Group (1 mentions)
Middlebrook 7h11 agar, by BD (1 mentions)
Adult bovine serum, by Thermo Fisher Scientific (1 mentions)
Amoxicillin, by Merck Group (1 mentions)
Fungizone, by Merck Group (1 mentions)
Middlebrook 7h10 agar, by BD (1 mentions)
Middlebrook 7h9 broth, by BD (1 mentions)
Glycerol, by Merck Group (1 mentions)
4 protocols using trimethoprim lactate
1
Intranasal M. bovis BCG Infection in Mice
2
Mycobacterial Burden Quantification
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Right lungs and spleens were harvested for CFU enumeration. Lung and spleen tissues were homogenized in 5 ml of sterile phosphate-buffered saline (PBS) and serially diluted on 7H10 agar (262710, BD Diagnostic) supplemented with 2.5 mg/L amphotericin B (A9528, Sigma), 26 mg/L polymyxin B sulfate (P4932-5MU, Sigma), 20 mg/L trimethoprim lactate (T0667-260mg, Sigma), 50 mg/L carbenicillin disodium (C3416-1G, Sigma), and OADC enrichment (R450605, Fischer) (25 (link)). CFUs were quantified following humidified incubation at 37°C for 2 to 3 weeks.
3
Selective Antibiotics and Bacterial Growth
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The effect of supplementing the media with selective antibiotics on the growth of bacteria on the media was examined by comparing bacterial growth on the media with the new medium not supplemented with antibiotics to the growth of the bacteria on media that had been supplemented with antibiotics after aerobic incubation at 37 C for 48 h. Modifications of two antibiotic mixtures used as selective Campylobacter media supplements were tested. One selective supplement tested was the CCDA supplement that contained 32 mg/L of cefoperazone and 10 mg/L of amphotericin B (Lyophilchem, Roseto degli Abruzzi, TE, Italy; CN 81037), with and without the addition of 10 mg/L or 20 mg/L of vancomycin hydrochloride (Sigma-Aldrich Co., St. Louis, MO, USA; CNs 75423). The other supplement was the Skirrow antibiotic supplement that contained 10 mg/L of vancomycin hydrochloride, 20 mg/L of trimethoprim lactate, and 2500 IU polymyxin B 2500 IU (Sigma-Aldrich Co., St. Louis, MO, USA; CNs T0667, and 1405-20-5, respectively). Concentrations listed for each supplement are the final concentration of antibiotics after addition to the medium.
4
Phenotypic Characterization of M. tuberculosis Isolates
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All M. tuberculosis M. tuberculosis M. tuberculosis Table 1 ): sterile lysed ovine blood prepared from sheep blood defibrinated (TSC Biosciences), adult bovine serum (Gibco), OADC and ADC enrichments (Becton Dickinson), while glycerol, malachite green, fungizone, polymyxin B, trimethoprim lactate and amoxicillin were all sourced from Sigma-Aldrich.
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