Bacterial Toxin Research Citations

Cells and Reagents:

…Polyclonal goat antibody (#771B) against Protective Antigen (PA) was from List Biological Laboratories (Campbell, CA) and used at 1/2000 dilution…

Cell Preparation, Reagents, and Stimulation:

In some experiments, cells were pretreated with Ac-YVAD-cmk (50 M, ALX-260028-M001, Enzo Life Sciences) for 30 min before S. Typhimurium infection or fliC transfection. Otherwise, cells were treated with anthrax protective antigen (1 g/mL, List Biologicals)…

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

… For the rPA, 7.5 g of rPA (LIST Biological Laboratories) was mixed with alum and either saline or 50 g of Poly I:C was added to the mix on the day of immunization. Antibody assays. To measure levels of antibodies to the rPA …

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

Animals and vaccinations:

For rabbit experiments employing Freunds adjuvant, Female New Zealand white (NZW) rabbits (Covance Research Products, Denver, PA) weighing approximately 2.5 kg were immunized on day 0 with a mixture of two MAPs (125 g per MAP) in CFA and boosted 4 times at two-week intervals with the MAPs in IFA. For studies using human-use adjuvants, NZW rabbits were injected with TB-LND(2) or PA83 (List Labs, Campbell, CA) according to the above schedule, …

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

Reagents:

Anthrax lethal factor (batch 1692A1B), and protective antigen (batch 17117A1B) from List Biological Laboratories.

Discussion:

In mouse macrophages of sensitive strains, LT triggers the formation of the NLRP1b inflammasome, culminating in pyroptosis and in the secretion of mature IL-1 and active caspase-1…

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

List Labs Product #169A (Anthrax Lethal Factor (LF-A), Recombinant from B. anthracis Native Sequence; Lyophilized) has been discontinued; however, Product #169L (Anthrax Lethal Factor (LF-A), Recombinant from B. anthracis Native Sequence; Liquid) is available for purchase.

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

Reagents:

…The SEB and protective antigen (PA; List Biological Laboratories, Campbell, CA, USA) were fluorescently tagged using an amine reactive DyLight 488 …

MMS sorting:

In target sorting, the streptavidin-depleted library was resuspended in 1ml PBS and was incubated with 600nM SEB-biotin for 45min. After incubation, the cells were centrifuged at 3000g for 5min and resuspended in 1ml PBS containing 110⁹T1 streptavidin beads. After 45-min incubation with the magnetic beads, the cells were loaded onto the MMS and separated using the MMS positive selection program. The bacterial cells selected as SEB positive binders by the MMS were plated in serial dilutions to determine the resultant library diversity, and the remaining cells were grown overnight in LB-Cm²⁵. In the second, third, and fourth rounds of sorting, the SEB concentration was decreased from 600nM in round one to 300nM in round two, 150nM in round three, and 75nM in round four. …

Off-Cell affinity and specificity:

After blocking, a 45 min binding step was performed with each protein dissolved in PBST and serially diluted across each well in the 96-well plate: SEB-HRP initial concentration 14 nM, Strep-HRP initial concentration 250 nM, HRP initial concentration 200 nM, and PA-HRP initial concentration 100 nM. …

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

IPEC-J2 cell culture conditions and bacterial adherence assays:

…To test for the effects of endogenous LT expressed by the bacteria on adherence in isogenic porcine-origin ETEC strains that had not demonstrably shown an effect of LT on colonization [14], inocula were prepared and directly applied to the IPEC-J2 cells, as described above. To test the effect of exogenous LT on adherence on these strains, LT (List Biological Laboratories) at a concentration of 1, 10, or 100 ng/ml per well was added (pre-incubation) 1 h prior to bacterial inoculation. To test the potential of GM1 ganglioside to mitigate the effects of LT on adherence, LT at a concentration of 10 ng/ml or 100 ng/ml was incubated with the cells 1 h prior to their inoculation with bacteria, and 10 ng/ml or 100 ng/ml GM1 ganglioside (Sigma) was applied to the IPEC-J2 cells at the time of bacterial inoculation. Three independent experiments were performed for each assay.

Test of effects of different carbohydrates on LT production and secretion:

To test the effects of different sugars on LT secretion, CAYE media containing glucose, sucrose or fructose at 0%, 0.25%, 0.5%, 1%, or 2% were prepared. Strains were cultured in different media as described above, with aliquots taken at 0, 2, 4, and 6 h post-inoculation (PI). These samples were centrifuged (2500g, 10 min), and supernatants taken to analyze for LT concentrations by GM1-ELISA as previously described [17]. Three independent experiments were performed for each assay. 

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

Female A/J mice were vaccinated with rPA (1 mg; List Laboratories) formulated with Al(OH) 3 (200 m g), SMIP-7.10 (25 m g), or both.

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

Peptides and proteins:

… Recombinant protective antigen (rPA) was obtained from List Biologicals (Campbell, CA ) ….

 Vaccines and immunization:

… For anthrax and influenza protein vaccines used in Chapter 3, lipids were prepared with S100 lecithin (Lipoid GmBH, Bermany).  These vaccines were delivered via intramuscular injections of 25 L on each the right and left leg.  Each 50 L dose of anthrax vaccine contained 1 g rPA and adjuvants as indicated in figure legends.  …

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

…MAPKKide was purchased from List Biological Laboratories (Campbell, …

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

• Product #530 – MAPKKide Peptide Substrate (o-Abz/Dnp) for Anthrax Lethal Factor
• Product #531 – MAPKKide Peptide Substrate (DABCYL/FITC) for Anthrax Lethal Factor