Citations

… (CFA) at a dose of 100 µg per mouse, followed by the administration of pertussis toxin (PT) (200 ng per mouse through intraperitoneal injection (IP); List biological …

Author did not specify which List Labs Pertussis Toxin was utilized. List Labs provides the following Pertussis Toxin products:
• Product #180 – Pertussis Toxin from B. pertussis, Lyophilized in Buffer
• Product #181 – Pertussis Toxin from B. pertussis, Lyophilized (Salt-Free)
• Product #179A – Pertussis Toxin from B. pertussis (in Glycerol)

… Then, 200 ng of pertussis toxin (List Biological Laboratories, Campbell, CA, USA) was intraperitoneally injected on days 0 and 2 after immunization …

Author did not specify which List Labs Pertussis Toxin was utilized. List Labs provides the following Pertussis Toxin products:
• Product #180 – Pertussis Toxin from B. pertussis, Lyophilized in Buffer
• Product #181 – Pertussis Toxin from B. pertussis, Lyophilized (Salt-Free)
• Product #179A – Pertussis Toxin from B. pertussis (in Glycerol)

… pertussis toxin (List Biological Laboratories, Campbell, CA);…

Author did not specify which List Labs Pertussis Toxin was utilized. List Labs provides the following Pertussis Toxin products:
• Product #180 – Pertussis Toxin from B. pertussis, Lyophilized in Buffer
• Product #181 – Pertussis Toxin from B. pertussis, Lyophilized (Salt-Free)
• Product #179A – Pertussis Toxin from B. pertussis (in Glycerol)

… Diphtheria Toxin List Biological Laboratories 150 …

• Product #150 – Diphtheria Toxin, Unnicked, from Corynebacterium diphtheriae

… After immunization, 0.25ug pertussis toxin (PTX) (List Biological Laboratories, Campbell, California, USA) dissolved in PBS was injected intraperitoneally on days 0 and 2. …

Author did not specify which List Labs Pertussis Toxin was utilized. List Labs provides the following Pertussis Toxin products:
• Product #180 – Pertussis Toxin from B. pertussis, Lyophilized in Buffer
• Product #181 – Pertussis Toxin from B. pertussis, Lyophilized (Salt-Free)
• Product #179A – Pertussis Toxin from B. pertussis (in Glycerol)

Toxin degradation assay

Toxin degradation was assessed using ADS024 CFS and reconstituted lyophilized cells. The CFS was prepared as previously described and was serially diluted 1:2 in BHI broth 10 times. Lyophilized ADS024 was reconstituted and serially diluted in phosphate-buffered saline. Purified C. difficile toxins A and B (0.2 mg/mL) were obtained from List Biological Laboratories (Campbell, CA, USA). Purified toxin was reconstituted in sterile water to contain 50 mM Tris, 50 mM NaCl, pH 7.5, and 0.1% trehalose and was stored at 2–8 °C; 2.5-μL (0.2 mg/mL) aliquots of purified C. difficile toxin A or B were co-incubated for 2 h at 37 °C with 30 μL of either ADS024 CFS, or lyophilized ADS024, and the reactions were stopped, heated, and electrophoresed in a 3–8% NuPAGE Tris–acetate protein gel as 15-μL volumes/well. The gel was electrophoresed at 150 V for 50 min. Western blot using TcdA or TcdB primary antibody (1:5000 dilution from List Biological Laboratories) and fluorescent secondary antibodies was performed.

• Product #152C – Toxin A from Clostridium difficile
• Product #155 – Toxin B from Clostridium difficile

EAE induction

All animal studies are approved by the Johns Hopkins University Animal Care and Use Committee. Animals received standard husbandry. C57Bl/6 mice (female, 6–10 weeks, Jackson Laboratories, n = 5–8) were injected s.c. with 200 μl of emulsion containing incomplete Freund’s adjuvant (Sigma F5506) Mycobacterium tuberculosis H37Ra (Difco BD 231141, 5 mg/ml) and myelin oligodendrocyte glycopeptide (MOG35-55, Johns Hopkins Synthesis and Sequencing Facility, 0.5 mg/ml). Mice were injected i.p. with 100 μl of pertussis toxin (List Laboratories #180, 300 ng) on the day of induction and 2 days later. …

• Product #180 – Pertussis Toxin from B. pertussis, Lyophilized in Buffer

… To deplete cDC1s in XCR1DTR mice, diphtheria toxin (DT; List Biological Laboratories) at 500 ng/mouse was injected intraperitoneally every three days beginning on day …

• Product #150 – Diphtheria Toxin, Unnicked, from Corynebacterium diphtheriae

… For imaging, cells were seeded in 96 well μClear® plates (Greiner Bio-One) or NuncTM MicroWellTM optical bottom 96-well plates (Thermo Fisher Scientific) and allowed to establish a confluent monolayer 16–24 h prior to evaluation by high content or scanning confocal microscopy, respectively. For immunofluorescence analysis, cells were seeded on a 12-mm microscope cover glass (VWR) in a 24-well plate overnight. Where indicated, BMMØs were cultured in the presence of either 10 ng/mL recombinant murine IL-4 or 100 U/mL recombinant murine IFN-γ (PeproTech) for 24 and 18 h respectively, prior to phagosomal assessment. Where indicated, cells were pulsed overnight with 2 mg/mL Bella Fluor (BF) 350SE-labelled (Setareh Biotech) dextran (70 kDa) with the dextran being chased into lysosomes the following day for 2 h. Where indicated, 2.5 mM of LLOMe was added to cells 30 min after phagocytosis of experimental particles and incubated for 2 h prior to imaging. Where indicated, 10 nM of myricetin (Alfa Aesar), 10 nM of kaempherol (Selleckchem) or 100 nM of EACC (Life Chemicals) were added to cells at the same time as experimental particles. Where indicated 0.5 μM Torin 1 (Tocris Bioscience), 0.5 μM rapamycin (Sigma Aldrich), 1× MEM amino acid solution (Sigma Aldrich), 10 ng/mL ultrapure lipopolysaccharide from S. Minnesota R595 (LPS; List Biologicals) were added to assay medium 30 min after the addition of reporter particles or 50 nM Lysotracker® Red (Invitrogen) 2 h after the addition of experimental particles and remained in the assay medium for the duration of phagosomal assessment.

• Product #434 – ULTRA PURE LPS from Salmonella minnesota R595 (Re)

… LPS from Salmonella Typhimurium was purchased from List Biological Laboratories …

• Product #225 – LPS from Salmonella typhimurium