Invivopure ph 7.0 dilution buffer

In T-cell transfer experiments, unfractionated CD4⁺ T cells were isolated from the spleens and lymph nodes of donor mice (WT, CD45.1, Stat1^−/−, Ifnar1^−/−Ifngr1^−/−) by negative selection (Miltenyi Biotec CD4⁺ T-cell isolation Kit, Cat No. 130-104-454). 1 × 10⁶ T cells (92.7–98.6% pure) were then adoptively transferred into recipient mice (Rag1^−/−, Il10rb^−/−Rag1^−/−) by i.p. injection in PBS unless otherwise stated. In some experiments, CD45.1⁺ T cells and Stat1^−/− T cells were mixed at a 1:1 ratio before being transferred into recipient mice (Rag1^−/−, Stat1^−/−Rag1^−/−). In some experiments, T cells were labeled with 5 μM CellTrace Violet (Thermo Fisher) in PBS + 0.1% FBS for 10 min at 37 °C prior to injection. All recipient mice were at least 6 weeks old and matched for sex, age, and housing between groups. Il10rb^−/−Rag1^−/− mice were monitored weekly for body weight changes post T cell transfer. For NK depletion assays, each mouse was first injected with 400 µg anti-NK1.1 (or isotype control) 1 day prior to T-cell transfer. For experiments lasting beyond 1 week, depletion of NK cells was maintained by injections of 200 µg anti-NK1.1 (or isotype control) at 1 and 2 weeks post transfer. All antibody injections were administered i.p in InVivoPure pH 7.0 Dilution Buffer (BioXCell).

Anti-p40 (Bio X Cell catalog number BE0051, RRID: AB_1107698) and mouse IgG2A isotype control (Bio X Cell catalog number BE0089, RRID: AB_1107769) monoclonal antibodies were diluted in InVivoPure pH 7.0 Dilution Buffer (Bio X Cell). Mice were injected intraperitoneally with 1 mg of antibody suspended in 200 μL buffer every 3 to 4 days for 3 weeks, beginning 14 days post H. hepaticus inoculation. The frequency of injections was determined based on preliminary studies. Mice without colitis received 200 μL of dilution buffer at each time point. Mice challenged with C. difficile continued to receive antibodies after infection until the end of experiment.

C57BL/6 mice were injected
sc with 100 μL of 0.25 × 10⁶ B16-OVA cells in
matrigel. When tumors reached a size between 50 and 100 mm³, mice were randomized according to tumor volume, and 0.4 ×
10⁶ OT-I T cells were adoptively transferred intravenously.
One day later, 200 μg of InVivoMab rat anti-mouse PD-1 antibody
(clone: RMP1-14, BioXCell, BE0146) was injected in InVivoPure pH 7.0
Dilution Buffer (BioXCell, IP0070) intraperitoneal in some mice, which
was given again on days 4 and 7. One day after OT-I T cell transfer,
mice either received 0.5 μg of pMHC^(SIIN) on IFs
or blank IFs in similar amounts as to the IF-pMHC^(SIIN), diluted in sterile PBS sc around the tumor. The weight and health
of mice was followed throughout the study. Tumor sizes were measured
every other day with a caliper. Tumor volumes were calculated as follows:
width × length × depth × 0.4. Tumor growth was followed,
and mice were sacrificed when the tumor reached the ≥1500 mm³ threshold. For the representation of tumor growth graphs,
tumor volumes of dead mice were kept at the threshold value after
euthanization until the end of the experiment. Tumors were formalin-fixed
(10% formalin) and paraffin-embedded (FFPE) for histological analysis.

To establish the lung metastasis model in Immunogenic mice, 4‐week‐old male C57 mice (n = 5 per group) were injected with hepa 1–6 WT or hepa 1–6 S100A9 cells (2 × 10⁶ cells suspended in 200 µL PBS through the tail veins. CD11b‐neutralizing antibodies (Bioxcell BE0007) were diluted to 1 µg µL⁻¹ concentration with InVivoPure pH 7.0 Dilution Buffer (Bioxcell IP0070) and administered i.p. at 100 µg per mouse and repeated once every 2 days.