Citations

2.10 Experimental autoimmune encephalomyelitis

For the induction of EAE, mice were immunized with MP4 antigen as previously described.36 In brief, male C57BL/6J mice at the age of 10 to 16 weeks were injected subcutaneously into both flanks with a total dose of 200 μg MP4 antigen (Alexion Pharmaceuticals, Cheshire, Connecticut), emulsified in complete Freund’s adjuvans (CFA). An intraperitoneal injection of 200 ng pertussis toxin (List Biological Laboratories, Hornby, Ontario, Canada; Cat #181) in PBS was given at the day of immunization and 48 hours later. Mice were sacrificed for analysis during the chronic stage of disease, 43 to 66 days after immunization.

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

… These solutions were prepared with 50 mM HEPES buffer with a pH of 7.4, on a 96-well plate, separately before being combined to result in final concentrations of 30 µM inhibitor, 150 nM enzyme, and 9 µM substrate (SNAPtide, List laboratories). An excitation wavelength of 490 nm was used and an emission at 523nm was observed and recorded for 5 mins. Two controls run were performed at the same time, containing no inhibitor. …

Author did not specify which List Labs SNAPtide was utilized. List Labs provides the following SNAPtide products:

• Product #520 – SNAPtide® Peptide Substrate (o-Abz/Dnp) for C. botulinum Type A Neurotoxin
• Product #521 – SNAPtide® Peptide Substrate (FITC/DABCYL) for C. botulinum Type A Neurotoxin
• Product #523 – SNAPtide® Peptide Substrate flP6(DABCYL/5-IAF) for C. botulinum Type A Neurotoxin

2.12. MOG/anti-CD3 challenge

An emulsion was prepared with 5 ​mg/mL MOG (New England Peptide, Gardner, MA) dissolved in PBS and 10 ​mg/mL heat-inactivated Mycobacterium tuberculosis H37Ra (Difco Labs, Detroit, MI) suspended in Incomplete Freunds Adjuvant (Difco Labs, Detroit, MI) at a 1:1 ratio (v/v). Female C57/BL6 mice were immunized with 100 ​μl total volume spread over 3 injection sites SC. Mice also received an ip injection of pertussis toxin (List Biological Labs, Campbell, CA) at 150 ng/mouse in 200 ​μl PBS.

2.14. In vitro T cell activation

An emulsion containing 100 ​μg PLP139-151 (New England Peptide, Gardner, MA) in incomplete Freund’s adjuvant (Difco Labs, Detroit, MI) supplemented with 200 ​μg ​M. tuberculosis H37Ra (Difco Labs, Detroit, MI) was administered to Balb/C mice subcutaneously over three sites in a total volume of 100 ​μl. Mice also received 100 ​ng pertussis toxin (List Biological Laboratories Inc, Campbel, CA) i.p. on day of 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)

2.2. Fabrication of human keratinocyte sheets

Normal Human Epidermal Keratinocytes (NHEK, Lonza, Basel, Switzerland) were expanded approximately ten times in cell number in keratinocyte culture medium (KCM) [8]. with mitomycin-C-treated 3T3 feeder layers as reported previously [9]. KCM was composed of the following components; Dulbecco’s modified Eagle’s medium (DMEM; Sigma Aldrich, MO, USA) and F-12 medium (Sigma Aldrich) were mixed at a 3:1 ratio、and supplemented with 5 μg/mL insulin (Eli Lilly, Indianapolis, USA), 10 ng/mL epidermal growth factor (EGF, Higeta Shoyu, Chiba, Japan), 1 nM cholera toxin (List Biological Laboratories, Campbell, USA), …

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

Mice and reagents.

C57BL/6 (B6), B6 CD45.1, BALB/c, CBA, B6 Foxp3-DTR, and B6 Foxp3-IRES-GFP mice were purchased from The Jackson Laboratory. As indicated, select lung and heart transplant recipients were treated with perioperative costimulatory blockade consisting of anti-CD40 ligand (250 μg intraperitoneally [i.p.], day 0) and CTLA-4–Ig (200 μg i.p., day 2) (Bio X Cell). Diphtheria toxin (DT) (List Biological Laboratories Inc.) (500 ng i.p. days 0, 1, and 7) was given to select B6 Foxp3-DTR heart transplant recipients to deplete Foxp3+ cells. …

• Product #150 – Diphtheria Toxin, Unnicked, from Corynebacterium diphtheriae

… Bacillus anthracis lethal factor (eg, List Biological Laboratories, 172B); …

• Prodcut #172 – Anthrax Lethal Factor (LF), Recombinant from B. anthracis

… 1A) Principle for isolating CT-binding bacteria. CT (List Biological Laboratories, USA) previously dialyzed against PBS was directly linked to Dynabeads M-270 Amine (Life technologies, UK) according to manufacturer’s instructions …

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

Ca2+ measurement

… To assess the role of G proteins in PAR2-mediated Ca2+ release, cells were pretreated either with PTX (100 ng/mL, 18 h; List Biological Laboratories) or YM-254890 (1 µM, 30 min; Wako Pure Chemical Industries) before agonist 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)

Histology and microscopy

For vermal Purkinje cell retrograde tracing, goat anti-cholera toxin B subunit primary antibody (1:15000, List labs, 703) and biotinylated horse anti-goat secondary antibody (1:2000, Vector, BA-9500) were used. …

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

… anthrax lethal factor (List Biological Labs #LL-169B) and protective antigen for anthrax lethal factor (LF) …

• Product #169L – Anthrax Lethal Factor (LF-A), Recombinant from B. anthracis Native Sequence
• Product #171E – Anthrax Protective Antigen (PA), Recombinant from B. anthracis