Bacterial Toxin Research Citations

EAE immunization:

The mice also received two 400 ng of intraperitoneal injections of pertussis toxin (List Biological Laboratories) in 100 µl of PBS immediately and 48 h later.

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)

Antibody variable region cloning and humanization:

The m1B7 and m11E6 variable region genes (17) were amplified from hybridoma cells by RT-PCR using degenerate primers and cloned into pAK100 as described (39). Positive clones were identified by monoclonal phage ELISA using a PTx-coated plate (1 mg/ml in PBS; List Labs), followed by sequencing.

ELISA and binding assays:

For indirect PTx ELISAs, a high-binding 96-well ELISA plate (Costar) was coated with 1 µg/ml PTx. The plate was blocked with milk for 1 hr, followed by incubation with duplicate anti-PTx antibody dilutions from 50 g/ml for 1 hr at 25°C. …

…PTx was injected in duplicate at 30 µL/min or 100 µL/min (m1B7, ch1B7), with concentrations between 5200nM diluted in running buffer (PBS or HBS, pH 7.4, 0.05% Tween). …

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)

Surgical procedures:

To visualize regenerating axons, cholera toxin subunit B (CTB; List Biological Laboratories) was injected using a protocol modified slightly from that in the previous report…After the sciatic nerve between the thigh muscles was exposed, a small incision was made on the perineurium just proximal to the trifurcation site. Four microliters of unconjugated CTB solution (0.1% in PBS) were slowly injected using the Hamilton syringe through the perineural incision and animals were killed 5 d after the injection.

p>Tissue processing and immunohistochemistry:

For immunohistochemistry, tissue sections were treated with 10% normal serum and 0.3% Triton X-100 for 1 h, and then primary antibodies dissolved in the same blocking solution were applied at 4°C overnight. The primary antibodies were…and goat anti-CTB (1:10,000; List Biological Laboratories).

• Product #104 – Cholera Toxin B Subunit (Choleragenoid) from Vibrio cholerae in Low Salt
• Product #703 – Anti-Cholera Toxin B Subunit (Goat)

Reagents:

… BoNT/B light chain were purchased from List Biological Laboratories (Campbell, CA, USA). …

Assay for BoNT Endopeptidase Activity:

The cleavage of the recombinant GST-Synaptobrevin-II protein substrate by the BoNT/B light chain domain was performed as suggested by the manufacturer (List Biological Laboratories Inc., Campbell, CA, USA) with minor modifications. Briefly, 10 M of GST-Synaptobrevin-II protein substrate was incubated with or without 60 nM of BoNT/B light chain and were then were treated with 2.5 g individual mAbs or a combination of antibodies for 3 h at 37 C (control samples were not pre-treated with mAbs) in a 10 L reaction (20 mM HEPES, pH 7.4, 1.25 mM DTT, 0.3 mM ZnCl₂ and 0.2% Tween-20). …

• Product #620A – Botulinum Neurotoxin Type B Light Chain, Recombinant

Cell culture and reagents:

… Protective antigen (PA), lethal factor (LF) and Escherichia coli derived LPS were purchased from List Biological Laboratories Inc. (California USA). The following is a list of antibodies (Abs) used in this study. …

Long-term TIR cells were generated as previously described [110].  Briefly, RAW 264.7 macrophages were treated with a cytolytic dose of LeTx (250 ng/mL LF and 500 ng/mL PA) for 5 h and surviving cells were plated in fresh culture medium.  After two weeks, surviving clones were individually picked and plated on a 96-well plate.  Each clone was tested for LeTx sensitivity and resistant clones were pooled and propagated. Short-term TIR cells were generated as previously described [110].  Briefly, RAW 264.7 macrophages were treated with a sub-lethal dose of LeTx (100 ng/mL LF and 100 ng/mL PA) for 5 h and then supplemented with fresh media overnight.  The next day surviving cells were pooled and plated onto new culture plates with fresh media.  BMDM-TIR cells were generated by Dr. Soon-Duck Ha as previously described [177].  Briefly, BMDMs were treated with a sub-lethal dose of LeTx (100 ng/mL LF and 100 ng/mL PA) for 24 h and surviving cells were plated onto new culture dishes with fresh media. …

Author did not indicate which specific lethal factor was utilized.  List Labs provides Product #172 (Anthrax Lethal Factor (LF), Recombinant from B. anthracis) and Product #169 (Anthrax Lethal Factor (LF-A), Recombinant from B. anthracis Native Sequence).

• Product #171 – Anthrax Protective Antigen (PA), Recombinant from B. anthracis

O s inibidor es Y – 27632 e Toxina A de Clostridium difficile fo ram obtido s da Calbiochem (San Diego, CA) e List Biological Labs (Campbell, CA, USA), respectivamente . …

• Product #152C – Toxin A from Clostridium difficile

Materials:

Standards of pure BoNT (150 kDa) were obtained from List Biological Laboratories, USA (BoNT/A) …

Experimental Procedures:

… To obtain high rates of successful dissection only the left phrenic nerve hemidiaphragms were excised from male mice of strain RjHan:NMRI … In each experiment, the preparation was first allowed to equilibrate for 15 min under control conditions. Then, the incubation solution was replaced by the toxin-containing solution (BoNT/A, B or E). After toxin application, the amplitude remained unchanged for some time (Figure 1), then decreased gradually depending on the toxin concentration. Toxin concentrations were such as to allow reduction of the contraction amplitude by 50% between 40 and 150 min. The times required to decrease the amplitude by 50% (paralysis time t_1/2 180 min) at different BoNT concentrations were used to construct the calibration curves for BoNT/A, B and E (Figure 2). Optimal power or logarithmic functions depending on the spread of the calibration curves were fitted to the calibration curves: y (BoNT/A; 0.031, 0.125, 0.5, 2, 8 pg/mL) = 46.387x^0.2622, …

Mice and immunizations

Eight‐ to ten‐week old C57BL/6 female mice were purchased from Japan SLC (Hamamatsu, Japan). For intranasal immunization of HBHA + CT, the mice were immunized with 30 μg of rHBHA and 1 μg of CT (List Biological Laboratories, Campbell, CA, USA) every 7 days, four times. Lymphocytes that had infiltrated the lungs were isolated and co‐cultured with APCs in the presence of HBHA. Two days later, cytokine concentrations in the culture supernatants were measured by ELISA. The cell cultures were performed in quadruplicate. For combined immunization, the mice were first s.c. immunized with 5 × 106 CFU of M. bovis BCG Tokyo strain (Japan BCG Laboratory) or PBS as a negative control. Twenty‐eight days later, the mice were intranasally immunized with 30 μg of rHBHA and 1 μg of CT or with PBS once a week four times….

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

-Galactosidase and CT assays:

… A positive-control assay for quantification of the level of CT in the samples was performed using purified CT (List Biological Laboratories).

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

Materials and Methods:

…CTB was visualized by incubating sections in a goat anti-CTB polyclonal antibody (List Biological Laboratories, 1 : 33 000 dilution) overnight, …

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