Citations

2.3. EAE induction

EAE (active EAE) was induced in 6–8-week-old female mice via subcutaneous immunization in the tail base bilaterally with 100 μg PLP-peptide in 100 μL of PBS in an emulsion mixed with 100 μL of incomplete Freund’s adjuvant (IFA; Difco, Detroit, USA) supplemented with 1 mg Mycobacterium tuberculosis H37Ra (Difco, Detroit, USA) as described previously [34]. The mice were then immediately injected intraperitoneally with 200 ng pertussis toxin (PT; List Biological Laboratories, Campbell, USA); the injection was repeated after 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)

Peanut Sensitization and Challenge

Peanut proteins were extracted from roasted defatted peanut flour (Golden Peanut, Alpharetta, GA) in PBS with 1 M NaCl, as previously described36. Female backcross mice, 4-6 weeks of age, were sensitized to peanut by oral gavage with peanut extract plus cholera toxin (List Biological Labs, Campbell, CA) once per week for four weeks. In the first three weeks, mice were given 2 mg peanut protein plus 10 μg cholera toxin, and in the fourth week, mice were given 5 mg peanut protein plus 10 μg cholera toxin. One week following the final sensitization, mice were bled via the submandibular junction, and serum was isolated for subsequent immunoglobulin quantification. …

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

EAE Induction and Treatment

The EAE model was induced as previously described [19]. Put simply, female C57BL/6 mice were subcutaneously immunized with 200 μl of 100 μg myelin oligodendrocyte glycoprotein (MOG35–55, Wuhan Haode Peptide, China) in incomplete Freund’s adjuvant (Sigma Aldrich, USA) containing 200 μg mycobacterium tuberculosis (strain H37Ra; Difco, USA) on day 0. Pertussis toxin (PTX; List Biologicals, USA) (200 ng) was administered intraperitoneally (i.p.) on day 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)

Materials And Methods

Primary human epithelial cells were obtained from three de-identified donor corneal rims courtesy of Dr.
Amy Estes from the Department of Ophthalmology, Medical College of Georgia, Augusta University, and The
Eye Guys, Eye Physicians and Surgeons of Augusta, GA. Primary corneal epithelial cell cultures were
established using an explant culture method [10]. Briefly, the corneal rim was visualized under a dissecting
microscope to confirm that it contained viable epithelial cells. If so, it was washed with Ca++ free PBS (pH
7.2), cut into 1-2 mm fragments, and placed epithelial side up in a 35 mm dish. After approximately 5
minutes in a laminar air-flow culture hood, corneal fragments became attached to the culture dish, and 1.5
mL of DMEM with 10% serum containing 40 μg/mL gentamicin, 1% ITS, and 100 ng/mL cholera toxin (LIST
Biological Laboratories, Inc., Campbell, CA) was added and the pieces were cultured in a humidified
incubator at 37°C with 5% CO2. The culture medium was replaced every two days. After 7 to 10 days, cells
were nearly 100% confluent. Cells were passaged using 0.25% trypsin, centrifuged at 500g for 5 minutes, and
subcultured in DMEM with 10% serum containing 40 μg/mL gentamicin (Life Technologies, NY), 1% ITS
(Fisher Scientific, Waltham, MA), and 100 ng/mL cholera toxin (LIST Biological Laboratories, Inc., Campbell,
CA). Cultured cells were inspected before use under the microscope to ensure minimal contamination by
stromal cells.

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

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/

EAE

EAE was induced in 8-12-week female C57Bl/6J/NOD/ShiLtJ mice using 150 μg of MOG35–55 (Genemed Synthesis, 110582) mixed with freshly prepared complete Freund’s Adjuvant (using 20 ml Incomplete Freund’s Adjuvant (BD Biosciences, #BD263910) mixed with 100 mg Myobacterium tuberculosis H37Ra (BD Biosciences, #231141)) at a ratio of 1:1 (v/v at a concentration of 5 mg/ml). All mice received two subcutaneous injections of 100 μl each of the MOG35-55/CFA mix. All mice then received a single intraperitoneal injection of 200 ng pertussis toxin (List Biological Laboratories, #180) at a concentration in 200 μl of PBS. Mice received a second pertussis toxin injection at the same concentration two days after EAE induction. …

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

… For cell ablation, 1 mg/ml stock solution of DTX (List Biological Laboratories, 150) was prepared with water, aliquoted and stored at -20°C until use. …

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

… Immunization was carried out at 0, 7, 14, 21 and 28 days. In the EAM-PTX (+) group, PTX (List Labs, cat. #: 181, Campbell, California, USA) 2 µg was intraperitoneally injected on days 0 and 7. …

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

… C. difficile toxin A (TcdA; List Biological Laboratories, Campbell, CA, USA). …

• Product #152C – Toxin A from Clostridium difficile

Pertussis toxin (PTx) List Biological Laboratories LBL-0901-80

In vivo experimental autoimmune uveitis (EAU) model

C57BL6/J mice were immunized via s.c. injection with 200 μl of emulsion containing 200 μg of IRBP1–20 in Complete Freund’s Adjuvant (CFA), and 0.2 μg of pertussis toxin was added i.p. on the same day. 100K-EVs were administrated via i.v. injection on days 0, 1, 2, and 7 and mice were sacrificed at day 18. After whole eyes were removed and fixed in 4% paraformaldehyde (PFA), sections were cut through the pupillary-optic nerve plane, and H&E staining was performed. …

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)