Citations

2.3.9. Immunohistochemistry for VGlut1

Following ex vivo MRI, spinal cords were washed in PBS, cryoprotected in 30% sucrose, frozen in OCT compound and cut in transverse 15 μm sections. Sections were immunostained using; rabbit anti-vGlut1 antibody (1:1000, #135302, Synaptic Systems), Goat-anti-CTB (1:2000, #703, List Biolgicals), Mouse-anti-NeuN (1:500, #MAB377, Millipore). Sections were counterstained with Alexa 546 and Alexa 488 (1:1000, #A10040 and #A10036, Thermofisher) and Dylight 650 (1:500, #ab96938, Abcam). Five sections throughout the L4-L6 segments were selected and traced α-motor neurons (Ctb+/ NeuN+) in the ventral horns were imaged at 400× magnification (LSM 710, Zeiss). Animals with fewer than five traced motor neurons were excluded from analysis.

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

Peptide-polymer synthesis and nanofiber preparation

For adjuvanted solutions, nucleotide adjuvants (cyclic di-AMP and CpG) were added just before fibrillization; all other adjuvants were added after fibrillization. Adjuvants were included in formulations at the following dosages per mouse: 10 μg of CTB subunit (List Labs, 104), …

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

Induction and evaluation of EAE

EAE was induced 35 days after γHV68 infection by injecting 100 μl of emulsified Complete Freund’s Adjuvant (DIFCO) containing 200 μg of MOG35–55 (GenScript) and 400 μg of desiccated Mycobacterium tuberculosis H37ra (DIFCO) subcutaneously. Mice also received two doses of 200 ng of pertussis toxin (List Biologicals) intraperitoneally at the time of immunization and 48 hours after induction. …

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)

… Mgl2-DTR mice were injected intraperitoneally with 1 µg of DT (List Biological Laboratories) 28 days after primary infection (loading dose). Maintenance dosing of 100 ng …

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

… The mucosal adjuvants used were CT (List Biological Laboratories, Campbell, CA, USA) …

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

Active EAE and adoptive transfer EAE

In active EAE model, … At 0 and 48 h after immunization, the mice were intraperitoneally treated with 500 ng of pertussis toxin (List Biological Laboratories, Inc., Campbell, CA, USA). …

In another EAE model by adoptive transfer, … At 0 and 48 h after immunization, the mice were intraperitoneally treated with 200 ng of pertussis toxin (List Biological Laboratories, Inc., Campbell, CA, USA). …

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)

2.3 MOG35–55 EAE immunization and administration

MOG35-55-induced the EAE in C57BL/6 mice was set up as described previously[20].Briefly, experimental mice were injected subcutaneously on day 0 with 150 µg MOG35-55 peptide emulsified in Freund’s complete adjuvant (CFA) containing
Mycobacterium tuberculosis strain H37Rv (2 mg/mL) (Difco Laboratories, Franklin Lakes, NJ, USA), then mice were injected intravenously with an additional 300 ng of pertussis toxin (PTX) (List Biological Laboratories Inc., Campbell, CA, USA) injection in 100 µl of PBS on day 0 and day 2 following sensitization. …

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)

Transplantation of ex vivo conditionally reprogrammed culture

Conditionally reprogrammed cultures (CRCs) of ASC and AC tumors isolated from KL;CC10 and KL;SPC mice were previously generated and were maintained in co-culture with γ-irradiated NIH3T3 fibroblasts in F12/DMEM media supplemented with 5% HI-FBS, 5 mg/ml insulin (I2643, Sigma-Aldrich), 24 mg/ml adenine (A2786, Sigma-Aldrich), 10 mM Y-27632 (ALX-270-M0055, Enzo Life Sciences), 10 ng/ml human recombinant epidermal growth factor (hr-EGF; 354052, BD Pharmigen), 0.4 mg/ml hydrocortisone (Sigma-Aldrich), 10 ng/ml cholera toxin (100B, List Biological Laboratories) and 1% penicillin/streptomycin (15140122, Thermo Fisher Scientific). 5-10×105 CRC cells were subcutaneously injected to the flanks of nude mice and the tumors were measured weekly with a caliper.

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

Antibodies were obtained from the indicated vendors: … and TcdB (List Bio, 754 A).

EAE

… 200 µg MOG35-55 peptide (Genemed Synthesis) in complete Freund’s adjuvant supplemented with 4 mg/ml Mycobacterium tuberculosis (BD Difco). Four hundred nanogram pertussis toxin (List Biological Laboratories) was administrated intraperitoneally at days 0 and 2 post-immunization. EAE symptoms were scored daily as follows: 0, asymptomatic; …

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)