Bacterial Toxin Research Citations

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)

2.2. Cell culture
The HDFa cell line was bought from the American Type Culture Collection (ATCC; Minnesota, USAHB-8065). It was cultured in Park Memorial Institute (RPMI -1640) culture media with glutamine 2 mM (bio west, Nampa, cat n L0498-500), 10% fetal bovine serum (PAA, Pasching Austria, cat. no. A11-151) and penicillin with streptomycin 1% (Lonza, Verviers, Belgium, cat. no. DE17-602E). The HDFa cells were grown in 50 cm2 flask (Greinerbio-one GmbH Maybachstr.272636 Frickenhausen, Germany) and preserved in typical humidified incubator supplied with 5% CO2, 95% air at 37 °C (New Brunswick Scientific- Innova co-170). Cells were washed with cold phosphate buffered saline, trypsinized, harvested and centrifuged to form cell pellets. The cultured HDFa were divided into four groups as follows:

Group I (Control): The HDFa cells received no treatment.

Group II (LPS): The HDFa cells were stimulated with LPS (10 ng/mL, Escherichia coli 055:B5, List Biological Laboratories, CA, USA) for 48 h (Skioldebrand et al., 2018).

• Product #203A – LPS from Escherichia coli O55:B5

EAE Model

EAE was induced in 8-wk-old female C57BL/6 mice by a subcutaneous injection with 100 µg of myelin oligodendrocyte glycoprotein (MOG) peptide (35–55aa: MEVGWYRSPFSRVVHLYRNGK) emulsified with Complete Freund’s Adjuvant on day 0 and an intravenous injection with 400 ng of pertussis toxin (List Biological Laboratories, Campbell, CA) 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)

STAR★Methods
Key resources table

Pertussis toxin List Biological Laboratories Cat#180

Mice, EAE induction and scoring
For EAE experiments, 6–8 weeks old mice were used. mSOD1 and 5xFAD mice were maintained up to 150 days and 15 months after birth, respectively. …

Mice, aged 6-9 weeks, were immunized subcutaneously with 100 μg MOG35-55 peptide (synthesized by Toray Research Center, Japan) emulsified in complete Freund’s adjuvant containing 1 mg heat-killed Mycobacterium tuberculosis H37RA (Difco). The mice were injected intraperitoneally (i.p.) with 100 ng of pertussis toxin (List Biological Laboratories, USA) at the time of immunization and 2 days later. …

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

… PBMCs were washed and stimulated with 1 µg/ mL Staphylococcus endotoxin B (SEB; List Biological Laboratories, California, USA) in the presence of Brefeldin A (Thermo Fisher Scientific, Massachusetts, USA) at 37°C and 5% CO2 for 16 hours …

… For induction of parental EAU, 8-12-week-old female and male mice were subcutaneously immunized at the base of the tail and both thighs with 500 μg human IRBP651-670 (LAQGAYRTAVDLESLASQLT, China Peptides Co., Ltd., Shanghai, China) peptide in a 200 μL emulsion in complete Freund’s adjuvant (CFA, St. Louis, MO, USA) containing 3.5 mg/mL Mycobacterium tuberculosis (H37RA, ATCC 25177), simultaneously with intraperitoneal injection of 1 μg pertussis toxin (PTX, List biological laboratories, inc., CA, 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)

Animals and induction of EAE

The EAE was actively induced in conventional inbred female C57Bl/6J mice (9-13 weeks old)
by immunization with myelin oligodendrocyte glycoprotein (MOG) 35-55 peptide (Prospec,
Rehovot, Israel) and complete Freund’s adjuvant (CFA) containing Mycobacterium
tuberculosis H37Ra (Sigma-Aldrich, Prague, Czech Republic). The MOG/CFA emulsion was
prepared by connecting two glass syringes to a 3-way connector and passing the solutions
through the connector. The emulsion was injected subcutaneously in two 50 μl doses. In total,
100 μg of MOG peptide per mouse was administered. To facilitate the transfer of lymphocytes
into the CNS, 300 ng of pertussis toxin (List Biologicals, Campbell, California, USA) in 200
μl of phosphate buffer saline (PBS) was injected intraperitoneally two hours and two days after
the EAE induction [14].

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)