Bacterial Toxin Research Citations

Mouse Model of EAE and Study Design

Animal experiments were conducted in accordance with national and institutional guidelines for the care and use of laboratory animals and with directive 2010/63/EU of the European Parliament and of the Council of 22 September 2010 and were approved by the Berlin State Office for Health and Social Affairs (LAGeSo, registration number G106/19). Experimental autoimmune encephalomyelitis was actively induced in 9- to 12-week-old SJL/J female mice (Janvier Labs, France; n = 29) by subcutaneous immunization with 250 μg proteolipid protein peptide (PLP139–151; purity 95%; Pepceuticals, Leicester, United Kingdom) and 800 μg Mycobacterium tuberculosis H37Ra (Difco, Franklin Lakes, NJ) diluted in 100 μL complete Freund’s adjuvant. In addition, 250 ng pertussis toxin (List Biological Laboratories, Campbell, CA) were injected intraperitoneally on day 0 (day of immunization) and day 2 of the study. …

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)

EAE induction and scoring.

Mice were immunized with CFA/MOG emulsion in the lower back on day 0. The emulsion was prepared by mixing MOG35-55 peptide (United Biosystems, catalog U104628) and CFA (MilliporeSigma, catalog F5881) with additional M. tuberculosis (BD Difco, catalog 231141; 200 μg/mouse). The mice also received an intraperitoneal injection of Pertussis toxin (PTx) (200 ng/mouse) (List Biological Technologies, catalog 180) on days 0 and 2. Unless otherwise noted, we induced EAE with this method, as Type-A EAE (100 μg MOG35-55/mouse and 200 μg M. tuberculosis/mouse) (10). In some experiments, Type-B EAE (10) was induced with CFA/MOG injection on days 0 and 7 (100 μg MOG35-55/mouse, 400 μg M. tuberculosis/mouse) and PTx (200 ng/mouse) on days 0, 2, and 7. EAE was scored as previously described (2, 3, 10).

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

… 200 ng of pertussis toxin (List Biological Laboratories, Inc., Campbell, CA, USA) was injected intraperitoneally (i.p.) on day 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)

… Non-induced age-matched littermate females were used as controls and were injected subcutaneously with an emulsion of PBS in CFA (Difco Laboratories, Detroit, USA), and with PTX (List Biological Laboratories, Inc., Campbell, CA, USA) at the same concentration and time points as the EAE animals …

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)

Induction and assessment of EAE

Myelin oligodendrocyte glycoprotein (MOG) 35-55 peptide was purchased from Genemed Synthesis, Inc (San Antonio, TX), was used as the immunogen. Rag1-/- mice (8-12 weeks old) received 5×106 CD4+ T cells from sex and age-matched C57BL6 mice or 1×106 CD4+ T cells from 2D2+CD24-/-MOG-/- mice [5]. The recipient mice were then immunized for the induction of EAE as we described [29]. Briefly, the T cell-recipient Rag1-/- mice were immunized subcutaneously with 200 µg MOG peptide in CFA (containing 400 µg of Mycobacterium tuberculosis) in a total volume of 100 µL. Mice were also injected with 150 ng of pertussis toxin (List Biological, Campbell, CA) in 200 µL PBS via the tail vein immediately after the immunization and again 48 hours later. EAE scores were evaluated on a scale of 0 to 5: 0, no clinical signs; 1, loss of tail tone; 2, wobbly gait; 3, hind limb paralysis; 4, moribund; and 5, death.

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)

… Cells were stimulated with ultrapure LPS from E.coli (O111:B4 serotype; List Biological Laboratories) at 10 or 100 ng/ml for up to 6 h …

• Product #421 – ULTRA PURE LPS from Escherichia coli O111:B4

EAE.
EAE was induced in female C57Bl/6J mice 8 to 12 weeks old using 150 μg of MOG35–55 (Genemed Synthesis, 110582) mixed with freshly prepared CFA (using 20 mL of incomplete Freund’s adjuvant [BD Biosciences, BD263910] mixed with 100 mg of Mycobacterium tuberculosis H-37Ra [BD Biosciences, 231141] at a ratio of 1:1 [vol/vol at a concentration of 5 mg/mL]). All mice received 2 s.c. injections of 100 μL each of the MOG/CFA mix. All mice then received a single i.p. injection of pertussis toxin (List Biological Laboratories, 180) at a concentration of 2 ng/μL in 200 μL of PBS. Mice received a second pertussis toxin injection at the same concentration 2 days after EAE induction. Mice were monitored and scored daily thereafter. EAE clinical scores were defined as follows: 0, no signs; 1, fully limp tail; 2, hind limb weakness; 3, hind limb paralysis; 4, forelimb paralysis; and 5, moribund.

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

… Pertussis toxin (List Biological Laboratories or MilliporeSigma) was dissolved in deionized water at 100 µg/ml, kept at 4 °C and freshly diluted to 2 µg/ml before use …

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 (PTX) was purchased from List Biological Laboratories, Inc. (USA) …

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)