Bacterial Toxin Research Citations

EAE Experiments

EAE was induced in male S1P1 -GFP signaling mice and GFP reporter mice aged 8-12 weeks. According to methods defined by Tsai et al [30, 31], mice were immunized with an emulsion containing 200 micrograms of Complete Freund’s Adjuvant (CFA), 100 micrograms of myelin oligodendrocyte glycoprotein (MOG35-55) peptide35–55, and 200 nanograms of Bordetella pertussis toxin (lot# 181236A1, List Biological Laboratories) on days zero and two. The spleen tissues were collected after eight days of immunization which marked the onset of the clinical EAE symptoms.

• Product #181 – Pertussis Toxin from B. pertussis, Lyophilized (Salt-Free)

Induction of the EAE model

To induce EAE, female mice at 6–8 weeks old (n = 14 and 17 wild-type and Smek1-/+ mice, respectively) were immunized along their back with MOG35–55 (2 mg/ml) and Mycobacterium tuberculosis (8 mg/ml) dissolved in complete Freund’s adjuvant. Immediately after immunization and 2 days after immunization, mice received an intraperitoneal injection of pertussis toxin (200 ng; List Biological, Campbell, CA) in 100 μL 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)

STAR★Methods
Key resources table

Cholera toxin List Biological Laboratory Cat#100B

• Product #100B – Cholera Toxin (AZIDE-FREE) from Vibrio cholerae

Cell preparation

In this work (1) SK-BR-3 (ATCC, Virginia, USA, HTB-30), HT-29 (ATCC, Virginia, USA), and CAKI-2 cells were thawed and cultured for two weeks in McCoy’s 5A (modified) Media (Wisent Inc., Quebec, Canada, 317-010-CL) supplemented with 10% fetal bovine serum (FBS), and 1% penicillin/streptomycin, (2) OCI-AML-5 cells (DSMZ, Braunschweig, Germany, ACC247) were thawed and cultured for two weeks in Alpha modified Eagle medium (Wisent Inc., Quebec, Canada, 310-010-CL) containing 10% FBS and 1% penicillin-streptomycin (Wisent Inc., Quebec, Canada, 450-201-EL) by volume, and (3) MCF-10A cells MCF-10A cells (Addex Bio, California, USA, C0006015) were thawed and cultured for three weeks in 1:1 Dulbecco’s Modified Essential Media and F12 media (ThermoFisher, Massachusetts, USA, 11330032), supplemented with 5% Horse serum (Sigma-Aldrich, Ontario, Canada, H1270), 20 ng/mL EGF (Peprotech, New Jersey, USA, AF-100-15), 0.5 ug/mL Hydrocortisone (Sigma-Aldrich, Ontario, Canada, H0888), 100 ng/mL Cholera Toxin (List Biological Laboratories, California, USA, 100B), …

• Product #100B – Cholera Toxin (AZIDE-FREE) from Vibrio cholerae

Experimental procedures

Since numerous commercial LPS preparations contain measurable contaminating proteins, for this study we utilized ultra-purified LPS (from Escherichia coli O111:B4) purchased from List Biological Laboratories, Inc. (Campbell, CA). Mice (C57BL/6: n = 5–8 animals; TLR4−/−: n = 4 animals for each experimental group) were injected with LPS solution (3 mg/kg, i.p.) prepared in sterile PBS (control). …

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

… Pertussis toxin (PTx) was from List Biological Laboratories …

Cytokine and Chemokine Production and Measurement

… To determine the inhibitory effect of PTx on cytokine and chemokine production, pretreatment with PTx (100 ng/mL, 16 h) was performed prior to any stimulation.

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)

Surgery, recordings, and tracer injections

… To precisely locate the left Su5, field potentials responding to electrical
stimulation of the left masseter nerve as well as to passive, sustained jaw-opening movements were
recorded. Then, the glass micropipette was changed to one filled with an anterograde tracer, biotinylated
dextranamine (BDA, 10,000 MW, Molecular Probes, Eugene, OR, USA) dissolved in saline in the first
experiment, or to one filled with a retrograde tracer, 1% cholera toxin B subunit (CTb; List Biological
Laboratories, Campbell, CA, USA) dissolved in 0.02 M phosphate-buffered saline (PBS, pH 7.4) in the
second experiment, in order to reinsert into the Su5. Signals recorded from the microelectrode were
amplified, filtered (300 Hz to 3 kHz), and stored in a computer at the sampling rate of 20 kHz (field
potentials) or 10 kHz (unit activity). Subsequently, BDA or CTb was extracellularly injected into the Su5
with electrophoresis (delivering 2.0-μA positive, 300 ms duration pulses, at 2 Hz, for 3–7 min). …

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

Cell Culture Conditions and Stimulations

… cells/well in a 96-well plate and stimulated for 24 h with one hundred µL undiluted sera from sham and burned mice. Salmonella minnesota R595 lipid A (List Biological Laboratories, INC Lot #: MLA-24A) was used as a positive control. …

• Product #401 – LIPID A monophosphoryl from Salmonella minnesota R595

In vivo injections

After recovery, each pigeon received unilateral injections of one tracer, BDA (10,000 molecular weight; Molecular Probes, Eugene, OR, USA) or CTB (low salt; List Biological Laboratories, Campbell, CA, USA) on the right hemisphere (Fig. 1a, 1g). BDA (10% in phosphate-buffered saline [PBS], pH 7.4) was used for anterograde labeling, whereas CTB (1% in PBS) was used for retrograde labeling. …

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

Cell surface binding assay

Binding of WT and mutated TcdB to cells was analyzed by the cell surface binding assay21. Briefly, cells were exposed to TcdB (10 nM) or TcdB mutants (10 nM) for 10 min at room temperature. Cells were washed three times with PBS and lysed with RIPA buffer (50 mM Tris, 1% NP40, 150 mM NaCl, 0.5% sodium deoxycholate, 0.1% SDS, with a protease inhibitor cocktail (Sigma-Aldrich). Cell lysates were centrifuged and supernatants were subjected to western blotting using chicken polyclonal anti-TcdB IgY (List Labs, #754A, 1:2000) and goat anti-chicken IgY H&L (HRP) (Abcam, ab97135, 1:2000) antibodies to examine the binding of TcdB mutants. Chicken polyclonal anti-actin antibody (Aves Labs, ACT-1010, 1:2000) was used for negative control.