29g insulin syringe, by BD - Product details

1

Cytokine-Loaded Hydrogel for Tumor Therapy

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B16F10 (10 [5 ]), LLC (2 × 10 [5 ]) or EL4 (10 [6 (link)]) cells were s.c. injected into the right flank of C57BL/6 mice. 4 T1 (5 × 10 [5 ]) or CT26 (5 × 10 [5 ]) cells were s.c. injected into the right flank of BALB/c mice. Treatment was initiated when the tumor reached ~6 mm in diameter.
In experiments using cells for treatment, 293 cells expressing IL12, GMCSF or IL2 were digested and resuspended in complete DMEM medium at a density of 1.5 × 10⁵ 293-IL12 + 1.5 × 10⁵ 293-GMCSF +1.5 × 10⁵ 293-IL2/45 μl. After the addition of 1.5 μl 2 M CaCl₂, 45 μl of 1.5% alginate (Sigma Aldrich, A0682) solution was added and mixed immediately to form hydrogel, which was slowly injected into tumors at 1.5 μl/mm² using a 29G insulin syringe (Becton).
In experiments using cytokine solution for treatment, 45 μl of solution containing 15 μl of each cytokine were mixed with 45 ul of 3% chitosan (Chitosan glutamate, Protosan G 213, NovaMatrix) solution. Next, the chitosan/cytokine solution was slowly injected into tumors at 1.5 μl/mm² using a 29G insulin syringe.
At the signs of disease progression, e.g., tumor size increase or new nodule appearance, additional injections were conducted until the tumor reached 12 mm in diameter.
A fifty-week-old BALB/c mouse bearing a 10 mm × 10 mm spontaneous tumor at ear root was intratumorally injected with 100 μl of chitosan/IL12 + GMCSF+IL2.

Zhang J., Jiang H, & Zhang H. (2018). In situ administration of cytokine combinations induces tumor regression in mice. EBioMedicine, 37, 38-46.

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2

Establishing a Rat Model of Ureteral Stenosis

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We established an original US animal model using microscopic vascular clamp placed on the left proximal ureter and subsequently removed 6 hours later. Rats were randomly divided into four groups. Sham rats had their left ureters exposed but not clamped via a flank incision. The remaining animals were subjected to establish US models. Under anaesthesia, the left ureter was visualized through a flank incision and clamped with sterile microscopic vascular clamp (15 mm; Mingmou Medical Apparatus Instruments, Suqian, Jiangsu, China) at upper ureter just below the lower pole of the left kidney (Figure 1A). For characterizing the pathogenesis of US causing by ureter damage, all the clamps were removed via the initial incision at 6 hours after surgery (Figure 1B). After clamps were withdrawn (not for Sham group), all rats received an injection in the left renal artery with either 100 μL PBS (Sham group, n = 8; US group, n = 8), 3.5 million MSCs in 100 μL PBS (MSCs group, n = 8), or 25 μg MSC‐EVs in 100 μL PBS (MSC‐EVs group, n = 8). We preferred to do the injection using a 29‐G insulin syringe (Becton, Dickinson and Company, USA) filled with 100 μL PBS/MSCs/MSC‐EVs.

Luo J., Zhao S., Wang J., Luo L., Li E., Zhu Z., Liu Y., Kang R, & Zhao Z. (2018). Bone marrow mesenchymal stem cells reduce ureteral stricture formation in a rat model via the paracrine effect of extracellular vesicles. Journal of Cellular and Molecular Medicine, 22(9), 4449-4459.

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3

ENTPD5 Knockdown and Overexpression Mice Models

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To generate ENTPD5-specific knockdown mice model, 20-week-old male db/db male mice and 8-week-old C57BL/6 male mice were injected adeno-associated virus expressing short hairpin ENTPD5 (AAV-sh-ENTPD5) through renal cortex multipoint injection subjected to B-mode ultrasound with the help of ultrasound doctors. For ENTPD5-specific overexpression mice model, 8-week-old C57BL/6 male mice were injected AAV8-ENTPD5 with the above operation. 100 μL of each AAV (1 × 10¹¹ pfu mL⁻¹) was aspirated with a 29 G insulin syringe (Becton Dickinson, USA). After injection AAV, 20-week-old db/db mice were fed on normal chow diets until to 28 weeks (Supplementary Table 4). Eight-week-old C57BL were fed on normal chow diets to 16 weeks and then performed the UUO surgery.

Xu L., Zhou Y., Wang G., Bo L., Jin B., Dai L., Lu Q., Cai X., Hu L., Liu L., Wu Y., Chang X., Huang Y., Song L., Zhang T., Wang Y., Xiao Y., Zhang F., Liu L., Shi M., Wang T, & Guo B. (2023). The UDPase ENTPD5 regulates ER stress-associated renal injury by mediating protein N-glycosylation. Cell Death & Disease, 14(2), 166.

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4

Chronic Pain Model in Rats

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Experiments were conducted in accordance with the National Institute of Neurological Disorders and Stroke/National Institute on Deafness and other Communication Disorders Animal Care and Use Committee and the International Association for the Study of Pain guidelines for the care and use of experimental animals. Male, Long Evans rats (Charles River) arrived at our facility at 6 weeks of age. Upon arrival rats were pair housed and each cage was allocated to either H6CTL or H3L6 diets (n=8 rats per group). Rats were maintained on an inverted, 12-hour light/dark cycle and were given ad libitum access to food and water. After 8 weeks of feeding, rats were subjected to a sensory testing battery. After 10 weeks of feeding, rats were injected intra-articularly at the ankle, with 50 μL of Complete Freund’s Adjuvant (Sigma-Aldrich, MO) using a 29G insulin syringe (Becton Dickinson, NJ).
The sensory testing battery was repeated 4 days and 20 days post injection. At 28 days post injection, relative hind paw weight bearing was evaluated, hind paw edema was measured with a caliper and rats were euthanized and brain, hind paw (ipsilateral and contralateral) and plasma were collected.

Domenichiello A.F., Wilhite B.C., Nara P., Pitcher M.H., Keyes G.S., Mannes A.J., Bushnell M.C, & Ramsden C.E. (2022). Biochemical and behavioral effects of decreasing dietary linoleic acid and increasing eicosapentaenoic acid and docosahexaenoic acid in a rat chronic monoarthrits model. Prostaglandins, leukotrienes, and essential fatty acids, 187, 102512.

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5

Cardiac ILC2 Transplantation in Mice

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CD45.2⁺Rag2^−/−γc^−/− mice were anesthetized with 3.5% isoflurane (Baxter) and tracheal intubation was performed immediately. While on intubation, mice were provided with oxygen with 2% isoflurane by mechanical ventilation system (Model 845, Harvard Apparatus). Pre-operational analgesics (0.05 mg/kg Buprenorphine, Reckitt Benckiser) and paralytics (1 mg/kg Succinylcholine, Henry Schein) were treated before operation. Thoracotomy was done to access the chest cavity and expose the ventricles of the heart. 5x10⁵ CD45.1⁺ cardiac ILC2s were injected into the myocardium on three ventricular locations using a 29G ½ insulin syringe (BD). Post-operational analgesics (0.05 mg/kg Buprenorphine) were administered during recovery.

Choi H.S., Won T., Hou X., Chen G., Bracamonte-Baran W., Talor M.V., Jurčová I., Szárszoi O., Čurnova L., Stříž I., Hooper J.E., Melenovský V, & Čiháková D. (2020). Innate Lymphoid Cells Play a Pathogenic Role in Pericarditis. Cell reports, 30(9), 2989-3003.e6.

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6

EAM Monocyte Transfer in Mice

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WT or IL-17Ra^–/– BALB/c mice on day 21 of EAM were depilated and anesthetized with 100% oxygen and 3.5% isoflurane (Baxter). Mice were subjected to an injection into the ophthalmic venous sinus. Roughly 4 – 5 × 10⁵ FACS sorted monocytes were injected with a 29G ½ insulin syringe (BD).

Hou X., Chen G., Bracamonte-Baran W., Choi H.S., Diny N.L., Sung J., Hughes D., Won T., Wood M.K., Talor M.V., Hackam D.J., Klingel K., Davogustto G., Taegtmeyer H., Coppens I., Barin J.G, & Čiháková D. (2019). The Cardiac Microenvironment Instructs Divergent Monocyte Fates and Functions in Myocarditis. Cell reports, 28(1), 172-189.e7.

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7

Purification and Labeling of Eosinophils

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Blood from donor IL-5Tg mice was collected in PBS with 100U/ml Heparin and overlaid on Histopaque-1119 (Sigma Aldrich) to remove red blood cells. Anti-CD90.2 and anti-CD45R(B220) microbeads (Miltenyi Biotec) were used to deplete lymphocytes and enrich eosinophils. Eosinophil purity (78%–95%) was determined by flow cytometry. For labeling, cells were stained with either CellTrace™ Violet (CTV) or CellTrace™ Far Red (CTFR) according to manufacturer’s instructions (Thermo Fisher). Recipient mice were anesthetized with 500 μL avertin or isoflurane and 1-8x10⁶ cells were injected to the mediastinal cavity or by retro-orbital injection using a 29G ½ insulin syringe (BD).

Choi H.S., Won T., Hou X., Chen G., Bracamonte-Baran W., Talor M.V., Jurčová I., Szárszoi O., Čurnova L., Stříž I., Hooper J.E., Melenovský V, & Čiháková D. (2020). Innate Lymphoid Cells Play a Pathogenic Role in Pericarditis. Cell reports, 30(9), 2989-3003.e6.

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8

EAM Monocyte Transfer in Mice

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WT or IL-17Ra^–/– BALB/c mice on day 21 of EAM were depilated and anesthetized with 100% oxygen and 3.5% isoflurane (Baxter). Mice were subjected to an injection into the ophthalmic venous sinus. Roughly 4 – 5 × 10⁵ FACS sorted monocytes were injected with a 29G ½ insulin syringe (BD).

Hou X., Chen G., Bracamonte-Baran W., Choi H.S., Diny N.L., Sung J., Hughes D., Won T., Wood M.K., Talor M.V., Hackam D.J., Klingel K., Davogustto G., Taegtmeyer H., Coppens I., Barin J.G, & Čiháková D. (2019). The Cardiac Microenvironment Instructs Divergent Monocyte Fates and Functions in Myocarditis. Cell reports, 28(1), 172-189.e7.

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9

Cardiac ILC2 Transplantation in Mice

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CD45.2⁺Rag2^−/−γc^−/− mice were anesthetized with 3.5% isoflurane (Baxter) and tracheal intubation was performed immediately. While on intubation, mice were provided with oxygen with 2% isoflurane by mechanical ventilation system (Model 845, Harvard Apparatus). Pre-operational analgesics (0.05 mg/kg Buprenorphine, Reckitt Benckiser) and paralytics (1 mg/kg Succinylcholine, Henry Schein) were treated before operation. Thoracotomy was done to access the chest cavity and expose the ventricles of the heart. 5x10⁵ CD45.1⁺ cardiac ILC2s were injected into the myocardium on three ventricular locations using a 29G ½ insulin syringe (BD). Post-operational analgesics (0.05 mg/kg Buprenorphine) were administered during recovery.

Choi H.S., Won T., Hou X., Chen G., Bracamonte-Baran W., Talor M.V., Jurčová I., Szárszoi O., Čurnova L., Stříž I., Hooper J.E., Melenovský V, & Čiháková D. (2020). Innate Lymphoid Cells Play a Pathogenic Role in Pericarditis. Cell reports, 30(9), 2989-3003.e6.

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10

Purification and Labeling of Eosinophils

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Blood from donor IL-5Tg mice was collected in PBS with 100U/ml Heparin and overlaid on Histopaque-1119 (Sigma Aldrich) to remove red blood cells. Anti-CD90.2 and anti-CD45R(B220) microbeads (Miltenyi Biotec) were used to deplete lymphocytes and enrich eosinophils. Eosinophil purity (78%–95%) was determined by flow cytometry. For labeling, cells were stained with either CellTrace™ Violet (CTV) or CellTrace™ Far Red (CTFR) according to manufacturer’s instructions (Thermo Fisher). Recipient mice were anesthetized with 500 μL avertin or isoflurane and 1-8x10⁶ cells were injected to the mediastinal cavity or by retro-orbital injection using a 29G ½ insulin syringe (BD).

Choi H.S., Won T., Hou X., Chen G., Bracamonte-Baran W., Talor M.V., Jurčová I., Szárszoi O., Čurnova L., Stříž I., Hooper J.E., Melenovský V, & Čiháková D. (2020). Innate Lymphoid Cells Play a Pathogenic Role in Pericarditis. Cell reports, 30(9), 2989-3003.e6.

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29g insulin syringe

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18 protocols using 29g insulin syringe

Cytokine-Loaded Hydrogel for Tumor Therapy

Establishing a Rat Model of Ureteral Stenosis

ENTPD5 Knockdown and Overexpression Mice Models

Chronic Pain Model in Rats

Cardiac ILC2 Transplantation in Mice

EAM Monocyte Transfer in Mice

Purification and Labeling of Eosinophils

EAM Monocyte Transfer in Mice

Cardiac ILC2 Transplantation in Mice

Purification and Labeling of Eosinophils

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