Bacterial Toxin Research Citations

… Pertussis toxin (List Biological Laboratories, Inc., Campbell, CA) was injected i.p. on the day of immunization and 48 h later. Weight and clinical scores were monitored every two days …

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)

Cell culture method

… For M1 polarized cells, on Day 3, PMA media is removed, and cells are washed with PBS. Thereafter, polarizing media consisting of complete media with LPS (List Biological Laboratories, 421, 100 ng/mL) and IFN-y (Sigma-Aldrich, SRP3093, 50 ng/mL) were added to the dish. The cells were incubated for another two days. On day 5, the media was removed, and cells were washed with PBS before fixing. …

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

MOG-induced EAE in / 6 mice

The mice were immunized by subcutaneous injections of 200 μg myelin oligodendrocyte glycoprotein (MOG) (35–55) (Genemed Synthesis, San Francisco, CA) in complete Freund’s adjuvant, containing 1 mg M. tuberculosis, strain H37RA (CFA; Difco, Detroit, MI), divided among two sites draining into the axillary lymph nodes. Sham mice, that served as negative controls, were injected with complete Freund’s adjuvant without MOG (35-55). After immunization (day 0) and on day 2, the mice received intraperitoneal injections of 200 ng pertussis toxin (List Biological Laboratories, Campbell, CA) dissolved in phosphate-buffered saline (PBS).

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)

Immunohistochemistry, image capture, and Imaris reconstruction

Mounted spinal cord sections were first incubated in PBS containing 0.1% Triton X (PBS-T) for 3 consecutive washes of 5 minutes each. Subsequently, spinal cord sections were incubated in 0.1% PBS-T solution containing primary antibodies and 10% heat-inactivated horse or donkey serum (Invitrogen) overnight at 4°C. Primary antibodies used were rabbit anti-DsRed (1:2000) [Clontech, 632496], guinea pig anti-vglut1 (1:1000) [Millipore, AB5905], goat anti-CTB (1:10 000) [List Biological Laboratories, 703], and chicken anti-parvalbumin (1:500) [sysy, 195006]. Following primary antibody incubation, spinal cord sections were washed with PBS for 15 mins (3x5min fresh solution). …

• Product #703 – Anti-Cholera Toxin B Subunit (Goat)

… A total of 2 × 107 cells were injected i.v. into naïve recipients, which then received 200 ng B. pertussis toxin (Ptx) (List Biological) i.p. 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)

LPS transfection and cell viability measurement

To induce pyroptosis in Hydra cells, dissociated Hydra cells were seeded into 12-well plates (approximately 107 cells per well) and stimulated with CTB (20 μg/μl; List Biological Laboratories) plus ultrapure LPS (1 μg/μl; E. coli O111:B4, InvivoGen). For inhibitor treatment, Hydra cells were pretreated with 20 μM Z-VAD-FMK (Sigma-Aldrich) for 30 min before adding CTB plus LPS. For propidium iodide (PI) staining, PI (5 ng/ml; Thermo Fisher Scientific) was added into medium to detect the loss of cell membrane integrity. The cytotoxicity induced by overexpression of HyGSDME wild type and its truncation mutants in HEK293T cells was determined by a CytoTox 96 assay kit (Promega) to detect the lactate dehydrogenase release. Static bright-field images of pyroptotic cells were captured by Nikon A1R microscope and processed by ImageJ. …

• Product #104 – Cholera Toxin B Subunit (Choleragenoid) from Vibrio cholerae in Low Salt

Immunizations

2W (EAWGALANWAVDSA) peptide was custom ordered from Genscript. 100 μg of peptide was prepared in a 50 μL emulsification of 1:1 (v/v) saline and complete Freund’s adjuvant (CFA)(Sigma) and injected subcutaneously at the base of the tail. For LPS treatments, 25–50 μg of E. coli derived lipopolysaccharide (List Biologicals) was prepared in 50 μL saline and administered intranasally into anesthetized mice. Peptide/CFA immunized or LPS-treated mice were harvested for tissues 12 days post-treatment unless indicated otherwise.

Author did not specify which List Labs LPS product was utilized in their research.
List Labs provides the following LPS products: https://www.listlabs.com/product-information/lipopolysaccharides/

Cartilage explant culture and experimental design

A previously described in vitro model12 was used to simulate the effects of OA on articular cartilage. Under aseptic conditions, harvested cartilage was cut into 5 X 5 X 1 mm explants totaling 36 explants per joint (72 total explants). The wet weight of the explants was recorded. Explants were placed into individual wells of 12-well tissue culture plates with 2 mL of DMEM, supplemented as described above, for a 48-hour equilibration period at 5% CO2 and 37 °C. After the equilibration period, culture media was replaced with the treatment culture media of 1 of the 12 respective treatment groups (Table 1). Half of the explants were challenged with equine recombinant interleukin 1 (rEq IL-1; 0.1 ng/mL) to simulate osteoarthritis, and the other half remained unchallenged to simulate healthy joint conditions. Explants were exposed to 1 of 6 concentrations of research-grade BoNT-A (0, 1, 10, 50, 100, or 500 pg/mL; List Labs). Explants remained in culture for 96 hours with a media change at 48 hours.

Botox injections

To induce temporary whisker paralysis, we performed Botox (Botulinum Neurotoxin Type A from Clostridium botulinum, List Labs
#130B) injections in the whisker pads of mice.13,44,45 Botox was reconstituted to a stock solution (40 ng/ml) with 1mg/ml bovine serum
albumin in distilled water. A microliter syringe (Hamilton Company) was used to inject 1 ml of a 10 pg/ml dilution made up from the stock
solution into the mystacial whisker pads of mice. Whisking ceased within 24 hours, and lasted for approximately one week before
whisker movements started to recover. A 50% supplemental dose of Botox was injected once per week, if necessary for additional
data collection. Botox injections (initial dose and supplements) were performed the day before imaging experiments, and absence of
whisking was verified before the start of each experiment.

EAE

Active immunization was induced by s.c. injection of MOG[35-55]11 (CHU de Québec) in incomplete Freund’s adjuvant (Difco) supplemented with 5 mg mL-1 Mycobacterium tuberculosis extract (Fisher). A dose of 25 μg MOG[35-55] per mouse was used to compare WT to Serpine1-KO mice, while 100 μg per mouse was used in EAE experiments involving Tim-3-Tg mice. In Figure 1D, passive EAE was induced by first immunizing Serpine1-WT 2D2 and Serpine1-KO 2D2 mice with MOG[35-55]. Nine days later, splenocytes were collected and stimulated ex vivo for 48 hours with MOG[35-55] (20 μg mL-1), rmIL-12 and rmIL-23. Next, 20×106 splenocytes were injected i.p. into unimmunized B6 recipients. Adoptive transfer of WT 2D2, Serpine1-KO:2D2 or RV-transduced 2D2 Th1 cells entailed i.v. injection (2×106) into Rag1-/- recipients at d5 of culture. In all EAE experiments, mice received 200 ng pertussis toxin (List Biological Labs) i.p. at d0 and d2. Mice were assessed for clinical symptoms daily as previously described (9).

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)