Bacterial Toxin Research Citations

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

… Immunohistochemical analysis was performed using biotin conjugated cholera toxin B subunit (List Biological Laboratories, Inc.) for detection of GM1 gangliosides, …

• Product #112 – Cholera Toxin B Subunit Biotin Conjugate

… After topical application of tetracaine hydrochloride to the cornea, they received a unilateral intraocular injection of cholera toxin subunit B (CTb) (List Biological Laboratories, Inc., Campbell, CA). …

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

… Twelve days after the optic nerve was crushed, the operated animals were anesthetized with a 7% chloral hydrate solution, and 3 µl 1% cholera toxin B subunit solution (CTB; List Biological Laboratories, Campbell, CA) was intravitreally injected. …

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

… Under aseptic conditions, the area to be injected was exposed, and rats were injected unilaterally with 2% FB plus 0.1% CTB (List Biological, Campbell, CA; four sites at 1.5 µl/site; …

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

… Mice were injected ip with pertussis toxin (PTX; List Biological, Campbell, CA) on days 0 and 2 (100, 300, 400 or 600 ng/mouse as indicated). Two types of control mice were included in these experiments: untreated mice and mice injected with PTX and CFA without PLP. …

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)

Animal care and use:

…Diphtheria toxin (DTX, List Biologicals) was injected as described previously [3], and does not affect sensory responses or body temperature when administered to wild-type mice, as previously described [3].

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

… E Coli O55 (list biological, Campell, CA) …

NOTE:  At the time this paper was written, Product #203 (5mg – LPS from Escherichia coli O55:B5) was utilized.

Product #203 (5 mg – LPS from Escherichia coli O55:B5) is no longer sold.

Product #203A (2.5 mg – LPS from Escherichia coli O55:B5) is available for purchase.

• Product #203A – LPS from Escherichia coli O55:B5

Mice and immunizations:

Mice were immunized with 1 (i.v. or i.n.) or 10µg (p.o.) CT (List Biological Laboratories, Campbell, CA), …

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

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