Bacterial Toxin Research Citations

FACS analysis

Differentiated hiPSCs were analyzed by FACS as described previously (Hayashi et al., 2017, 2018). Cells were then dissociated using StemPro™ Accutase™ Cell Dissociation Reagent (Thermo Fisher Scientific) and resuspended in cold KCM medium (DMEM without glutamine and Nutrient Mixture F-12 Ham [3:1; Thermo Fisher Scientific] supplemented with 5% FBS [Japan Bio Serum, Hiroshima, Japan], 0.4 μg/mL hydrocortisone succinate [Wako], 2 nM 3,3′,5-triiodo-l-thyronine sodium salt [MP Biomedicals, Santa Ana, CA, USA], 1 nM cholera toxin [List Biological Laboratory, Campbell, CA, USA], 2.25 μg/mL bovine transferrin HOLO form [Thermo Fisher Scientific], 2 mM l-glutamine, 0.5% insulin transferrin selenium solution [Thermo Fisher Scientific], and 1% penicillin-streptomycin solution). …

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

A total of 1 µL of 1% CTB (List Biological Labs, Campbell, CA, USA) solution was slowly injected under the epineurium of the sciatic nerve on the left leg using a 10 µL micro-syringe (Hamilton Company). Following injection, the incision was sutured closed using …

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

ELISA

Schistosoma mansoni egg antigens (SEA) and tetanus and diphtheria-specific IgG1 endpoint titers were determined by enzyme-Linked immunosorbent assay (ELISA) using the mAb X56 (BD) and Immulon 4HB plates (Thermo Fisher Scientific) as previously described [21]. Briefly, plates were coated with 2μg/mL tetanus (List Labs), diphtheria (Sigma), or SEA. The following morning plates were blocked with 1% milk and incubated with serial dilutions of primary antibody, followed by incubation with anti-mouse IgG1 ads-HRP antibody (Southern Biotech) and ABTS substrate. Plates were read at 405 nm at room temperature on a BioTek plate reader.

• Product #190 – Tetanus Toxin from Clostridium tetani

… For the detection of any GM1-binding CTB molecules irrespective of the length or sequence of C-terminal extension, a previously established method using a goat polyclonal anti-CTB antiserum (List Biological Laboratories, Campbell, CA) was employed …

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

2.8 Adoptive transfer

For passive EAE, mice were euthanized 10 days after MOG35–55 immunization, and spleen and lymph nodes were collected as previously described [27]. Single cells were incubated for 3 days in the presence of 25 µg/mL MOG35–55, 10 ng/mL recombinant murine IL-23 (R&D, 1887-ML-010), and 2 ng/mL IL-2 (R&D, 402-ML-020) at 1 × 107 cells/mL. CD4+ T cells were purified by CD4+ T cell isolation kit and 1 × 106 cells per mouse were transferred via intravenous injection. Pertussis toxin (200 ng/mouse, List Biological Laboratories, 181) was injected intraperitoneally (i.p.) on days 0 and 2. EAE score was assessed by daily recording using a 0–5 scale.

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

Experimental design of EAE mice

Experimental autoimmune encephalomyelitis (EAE) was induced in mice by subcutaneous immunization (on Day 0) with 50 μg of the myelin oligodendrocyte glycoprotein 35-55 peptide (MOG35-55; Peptide International, Louisville, KY, USA) emulsified in complete Freund’s adjuvant (Difco Laboratories, Detroit, MI, USA) supplemented with Mycobacterium tuberculosis extract H37Ra (Difco Laboratories). In addition, mice received 250 ng pertussis toxin (List Biological Laboratories, Campbell, CA, USA) intravenously on Day 0 and intraperitoneally on Day 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)

EAE experiment, histology, and immunofluorescence.

Immunization of female C57BL/6 mice (The Jackson Laboratory, Bar Harbor, ME, ages 8-10 weeks) with 200 µg of myelin oligodendrocyte glycoprotein (MOG) 35-55 (PolyPeptide Laboratories, San Diego, CA) in complete Freund’s adjuvant containing 400 µg of Mycobacterium tuberculosis per mouse (subcutaneous injection of 0.2ml volume), followed by pertussis toxin (List Biological labs Inc), was administered via intraperitoneal (i.p.) injection at day 0 (75ng per mouse) and day 2 (200ng per mouse) 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)

… Free-floating sections were washed in 0.01 M PBS, followed by incubation in hydrogen peroxide for 20 min and incubated for 24 h at room temperature (RT) with a goat anti-CT-B antibody (1:10,000 List Biological Laboratories, CA, USA) …

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

… In addition, the mice were administrated 200 ng of Pertussis toxin (List Biological Laboratories) intra-peritoneally (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)

Induction and treatment of EAE

To induce EAE, we injected mice (C57BL/6aged 8-10 weeks) with myelin oligodendrocyte glycoprotein (MOG residues35–55). The sequence of peptide was Met-Glu-Val-Gly-Trp-Tyr-Arg-Ser-Pro-Phe-Ser-Arg-ValVal-His-Leu-Tyr-Arg-Asn-Gly-Lys, and the peptide purity was more than 95 % (CL. Bio-Scientic Co., LTD., Xi’an, China). We admixed 100μg MOG35–55 peptide and complete Freund’s adjuvant containing 5mg/ml of heat-killed H37Ra, a Mycobacterium tuberculosis strain (Difco Laboratories). And then the mixture was injected into mice subcutaneously. Pertussis toxin (400ng) (List Biological Laboratories) in PBS and NaCl (50mM) was administered i.p. subsequently and the second time after 24 h [25]. …

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)