Bacterial Toxin Research Citations

Immunization, Nb infection and in vivo treatment

All immunization procedures were conducted in the rear footpad with 20 µL injection volume per footpad. Mice were immunized as indicated in each Figure with 5 µg low-endotoxin OVA (Worthington Biochemical Corporation) mixed with either 50 µg papain (P4762, Sigma), 10 µL FCA (F5881, Sigma), 10 µL alum (77161, Thermo Scientific), or 10 µg CpG2216 (Invivogen) in phosphate-buffered saline (PBS). For Nb infection, mice were injected subcutaneously in the right ear with 600 third-stage (L3) larvae (gift from William Gause, Rutgers New Jersey Medical School) in 0.2 mL of PBS. For DC depletion, mice were injected i.p. with 500 ng DT in PBS (List Biological Laboratories) at indicated time-points. …

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

Preparation of DT-conjugated latex beads and ICV injection

Two microliters of 1 μm carboxylated latex beads (Sigma-Aldrich; L4655) were diluted to 40 μl in ACSF (artificial cerebrospinal fluid)(Coolaber, SL6630-500ml), followed by co-incubation with 160 μl ACSF containing 100 μg of DT (List labs; 150) in a light-protected tube with continuous vortex overnight at room temperature. After that, beads were washed with cold PBS for three times to remove unbound free DT. Finally, the DT-conjugated beads were resuspended in 40 μl PBS. BSA-conjugated beads were used as controls. Beads were injected ICV at a volume of 5 μl per mouse at 1 μl min−1.

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

Treg cell transfer, diphtheria toxin, BrdU, and anti-amphiregulin administration

Thymi and spleen from 6–8-week-old female Foxp3RFP or Foxp3RFPRag2GFP were harvested and strained through a sterile 40 μm filter. Next, red blood cells were lysed and stained before sorting on a 70micron nozzle. 100.000 RFP+ thymic or splenic Treg cells or Rag2GFP+ or Rag2GFP- Thymic Treg cells were then injected retro-orbitally into mice 3 hours after SL-TBI. Diptheria toxin (DT; List Biological Laboratories) was administered at a dose of 50 ng/g in PBS via IP injections on 2 consecutive days. …

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

2.4. Evaluation of Anti-Diphtheria Toxin Antibodies

Firstly, 96-well plates (Falcon MicroTest 111; Becton Dickinson, Oxnard, CA, USA) were coated with 1 pg of diphtheria toxin antigen (List Biological Laboratories, Campbell, CA, USA) per well and diluted in 50 µL of phosphate-buffered saline (PBS). …

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

Mouse Treatment

For depletion of macrophages, 1 day before tumor inoculation, mice were injected via the tail vein with 200 µL mouse−1 clodronate liposomes or control PBS liposomes (40337ES10 or 40338ES10, Yeasen Biotechnology) every 4 days. For depletion of CD8+ T cells, mice were injected intraperitoneally with anti-mouse CD8α antibody (12.5 mg kg−1, BE0061, BioXcell) or isotype antibody (BE0090, BioXcell) every 3 days after tumor inoculation. For depletion of Tregs, 2 days after tumor implantation, C57BL/6J Foxp3DTR mice were performed by intraperitoneal injection of diphtheria toxin (12.5 µg kg−1, 15047A1, List Labs) or PBS every 3 days. For neutralization of CCL20 in vivo, tumor-bearing mice were injected intraperitoneally with 10µg dose−1 anti-mouse CCL20 antibody (AF760, R&D Systems) or isotype every 3 days from the third day. …

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

White blood cell profile detection

For inducible ablation of CD26+ cells at the injury sites, each mouse was locally injected with 50 ng diphtheria toxin ((List Labs) daily for 5 days. At day 3 after last time of DTx injection, peripheral blood from PBS and DTx group were collected. White blood cell profiles were detected with auto hematology analyzer (BC-5000 Vet) according to operation manual.

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

Neuronal Ablation

Mice carrying the En1Cre;hCdx2::FlpO, Chx10Cre;hCdx2::FlpO, Hes2iCre;hCdx2::FlpO, En1Cre;Chx10Cre;hCdx2::FlpO alleles in addition to the effector allele Tauds-DTR received i.p. injections of diphtheria toxin (DTx, 50 ng/gram of weight; List Biological Laboratories) at P28 and P31. Hes2iCre;VGATFlpO;Tauds-DTR mice and littermate controls Hes2iCre;Tauds-DTR mice received intrathecal injections of diphtheria toxin (DTx, 10 ng; List Biological Laboratories) at P28, P30 and P32. FlpO negative mice injected with DT were used as controls.

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

… mice were injected intraperitoneally with 500 ng boluses of un-nicked diphtheria toxin (DT) (150, List Biologics), every other day. …

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

Animals

… Male, Advillin-Cre+/−, mice were crossed with female, CalcaWT/GFP (Figure 1D), mice to generate CalcaWT/GFP, CalcaWT/DTR (Figure 1C), or wildtype CalcaWT/WT (Figure 1A) offspring. Diphtheria toxin (DTX; List Biological Laboratories, Product #150) was reconstituted to 20 μg/mL in sterile 0.9% saline, aliquoted, and stored at −80°C until used for injections. Once animals reached 10–12 months of age, DTX was administered by i.p. injection (100 μg/kg) twice with 72 h between the two injections. Animals were sacrificed and prostate tissue was harvested 14 days after the second injection (Figure 1E).

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

… Unnicked DT from Corynebacterium diphtheriae was purchased from List Biological Laboratories. Zbtb46-DTR bone marrow chimeras were injected i.p. with a first dose of 640 ng DT per mouse, …

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