Citations

… White quinoa seeds (originating from Peru) were purchased from Procudan (Kolding, Denmark) and ground to flour. Capsaicin 97% pure (M2028, Sigma-Aldrich, St. Louis, MO, US) and CT (List Biological Laboratories, Campbell, CA, US) were included as control substances. …

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

… Pertussis toxin (#180) was from List Biological Laboratories. …

Induction and assessment of EAE

To induce type A EAE, MOG35-55 peptide (300 µg per mouse) was emulsified with complete Freund’s adjuvant (200 µg M. tuberculosis H37Ra [Mtb] per mouse), and then subcutaneously injected in the flanks and neck of mice on day 0 for one immunization. Pertussis toxin (250 ng per mouse) was applied intravenously on days 0 and 2 after immunization. …

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

2.2. Antigens and antibodies
TeNT was obtained from List Biological Laboratories (USA).

2.3. Phage display selection
Llamas were injected intramuscularly simultaneously with both a ready to use tetanus toxoid vaccine intended for use in horses (1 ml, MSD Animal Health) and rTTC at day 0, 21 and 42. The rTTC (8 µg per injection) was emulsified with either Stimune adjuvant (Thermofisher Scientific, the Netherlands) for day 0 and 21 or with IMS1312 adjuvant (Seppic, France) for day 42. Serum samples were taken at day 0, 28 and 49. Heparinized blood samples were taken at day 28 and 49 for subsequent generation of phage display immune libraries in phagemid vector pRL144 [37], [41]. The libraries were rescued using VCSM13 helper phage. Phage display selections were performed by biopanning [42] with TeNT or rTTC presented by passive adsorption. Alternatively, plates were coated with cAb-TT2 or mAb TT10 [28], [39] in 50 mM NaHCO3 buffer (pH 9.6) for presentation of captured of TeNT. …

2.4.1. Analysis of E. coli-produced VHHs
Individual E. coli-produced VHHs clones were screened in an ELISA with coated TeNT or rTTC (at 1 µg/ml and 4 µg/ml respectively), and with cAb-TT2 or mAb TT10 (both were coated at 1 µg/ml in 50 mM NaHCO3 buffer (pH 9.6)) immobilised TeNT (at 1 µg/ml). …

2.4.2. GT1b-TeNT inhibition ELISA
A GT1b-TeNT inhibition ELISA was performed as reported [15]. Briefly, plates were coated with 10 µg/ml GT1b ganglioside in methanol at 100 µl/well by overnight incubation at RT. All subsequent incubations were performed in PBS containing 0.5% BSA and 0.05% Tween-20 for 1 h at RT. TeNT (1 µg/ml) was preincubated with VHHs, E. coli culture supernatants or mAbs in 100 µl/well in a plate for 1 h at RT. …

• Product #190 – Tetanus Toxin from Clostridium tetani

… Mice also received intraperitoneal (ip) injections of 200 ng pertussis toxin (List Biological Laboratories, Campbell, CA) on day 0 and day 2, with the GP group receiving a GP: MOG 35-55 boost on day 4 post 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)

Mice and EAE induction

… Wild-type littermate controls with same genetic background were used. All mice were maintained in specific pathogen-free conditions in the Laboratory Animal Department of Central South University. … EAE was induced by the use of myelin oligodendrocyte glycoprotein (MOG35–55, H-MEVGWYRSPFSRVVHLYRNGK-OH, GL Biochem) in 8-week-old female mice. Each mouse was immunized by subcutaneously injection with 200 μg of MOG35–55 emulsified in Complete Freund’s Adjuvant (CFA, Sigma) with 500 μg mycobacterium tuberculosis (strain H37RA, DIFCO). Mice were then intraperitoneally injected with 200 ng of pertussis toxin (List Biological Laboratories) at the time of immunization and 2 days after. …

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)

In vivo murine experiments

BALB/c 6- to 8-week old female mice (Charles River Laboratories, Inc., Worcester, MA) were used for the in vivo experiments. Mice were nasally vaccinated on days 0, 14, and 28 with 20 or 40 μg of exosomes derived from RAW264.7 cells infected with wild-type UK-1 S. Typhimurium (χ3761; exosomes were collected at two times points, 24–48 hpi, and pooled, n = 8 for each sample type). In some animals, we also included 2.5 μg of cholera toxin (CT; List Biologicals) or MPLA as an adjuvant (n = 8). …

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

… pertussis toxin (List Biological Laboratories, Campbell, CA) was injected i.p. at day 0 and day 2. In SJL/J relapsing–remitting EAE experiments, 9-wk-old SJL/J female mice were induced to EAE as described above using …

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)

… Mice were injected i.p. with 20 µg/kg of pertussis toxin (PTX) (List Biological Laboratories) immediately after and 2 d postimmunization …

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 Mice and cells

,,, Loosely adherent bone marrow derived dendritic cells (BMDCs) were collected after seven days. BMDCs were incubated 36 hours with 10 ng/mL Ultrapure Salmonella minnesota LPS (List Biologicals Laboratories, Campbell, CA) to generate activated DCs.

• Product #434 – ULTRA PURE LPS from Salmonella minnesota R595 (Re)

Induction and Assessment of EAE

EAE was induced in 8–12-week-old female BCMA−/− mice along with age- and sex-matched BCMA+/+ control mice. Mice were immunized with 150 μg MOG35–55 (Genemed Synthesis Inc., San Antonio, TX) emulsified in complete Freund adjuvant (1.5 mg/mL heat-killed Mycobacterium tuberculosis), followed by an IP injection of 250 ng Bordetella pertussis toxin (List Biological Labs, Inc., Campbell, CA) in phosphate-buffered saline (PBS) at the time of and 2 days 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)