Human lung microvascular endothelial cells were treated with a pool of target-specific 19–25 nt siRNAs designed to knock down either ENaC-α conducting subunit or ASIC1a gene expression, and non-specific, non-targeting siRNA were obtained from Ambion (Grand Island, NY, USA). All siRNA’s were received in lyophilized form. HL-MVEC were transfected at 70–80% confluence with 50–75 nM final concentration of siRNA using siPORT™ Amine transfection reagent (Ambion, Life Technologies, Grand Island, NY, USA) and used for further experiments at 48 h post transfection.
Siport amine transfection reagent
Manufactured by Thermo Fisher Scientific
Sourced in United States
The SiPORT Amine transfection reagent is a laboratory product designed to facilitate the delivery of nucleic acids, such as DNA or RNA, into cells. It is a formulation of cationic lipids that can form complexes with the genetic material, enabling its efficient uptake by the target cells. The SiPORT Amine transfection reagent is intended for use in a variety of cell types and experimental applications that require the introduction of foreign genetic material into cells.
Automatically generated - may contain errors
Lab products found in correlation
Lipofectamine 2000 transfection reagent, by Thermo Fisher Scientific (2 mentions)
Matrigel, by BD (1 mentions)
Si nc, by Thermo Fisher Scientific (1 mentions)
Si erbb2, by Thermo Fisher Scientific (1 mentions)
Sirna constructs, by Thermo Fisher Scientific (1 mentions)
Opti mem reduced serum media, by Thermo Fisher Scientific (1 mentions)
Amaxa nucleofection, by Lonza (1 mentions)
Haecs, by Thermo Fisher Scientific (1 mentions)
Ebm 2 medium, by Lonza (1 mentions)
Rpmi 1640 medium, by Merck Group (1 mentions)
20 protocols using siport amine transfection reagent
1
Knockdown of ENaC-α and ASIC1a in Human Lung Endothelial Cells
2
Anti-tumor effect of miR-720 silencing
3
Prostate Cancer Mouse Xenograft Model
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Five castrated nude mice (4-week-old; Institute of Zoology, Chinese Academy of Sciences) for each group received subcutaneous injections of 1 × 107 CS-FBS-treated DU145 or 5 × 106 CS-FBS-treated C4–2 cells, mixed with 1× Matrigel (BD Bioscience) in a volume of 100 μL, in both lateral flanks. Once palpable tumors developed, tumor width and length were measured twice a week using calipers. When the tumors reached an average volume of 150 mm3, 6.25 mg Ambion® miRNA mimics (miR-708/miR-C) complexed with 1.6 mL siPORT Amine transfection reagent (Ambion) or 50 mg/kG LY294002 in 50 μL phosphate-buffered saline was delivered intratumorally at 4-day intervals. The dosage was selected based on previous results18 (link). Mice were euthanized 2 days after the last treatment (day 58).
4
Silencing Estrogen Receptor Expression
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siRNAs used herein were as follows: ER-siRNA: Sense 5′-CAGGCACAUGAGUA ACAAATT-3′ and antisense 5′-UUUGUUACUCAUGUGCCUGAT-3′ (Ambion/Life Technol-ogies, Washington, DC); and negative control-siRNA (AM4461, Ambion). Subconfluent proliferating cells were treated with ER- or control-siRNA (100 nM) using siPORT Amine transfection reagent (Ambion). Western blotting revealed that a minimum of 48-hr exposure to 100 nM of ER-siRNA was required to obtain >75% reduction of ER.
5
Silencing PKAα and PKC-α in HL-MVEC
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HL-MVEC were treated with a pool of 3 target-specific 19–25 nt siRNAs designed to knock down either PKAα catalytic subunit or PKC-α gene expression. These were obtained from Santa Cruz Biotechnology, Inc. (Dallas, TX, USA). Also a non-specific, non-targeting siRNA was ordered from the same manufacturer. All siRNA's were received in lyophilized form. HL-MVEC were transfected at 70–80% confluence with 75 nM final concentration of siRNA using siPORT™ Amine transfection reagent (Ambion, Life Technologies, Grand Island, NY) and used for further experiments at 48 h post transfection.
6
Silencing NF-κB Pathway Inhibits Tumor Growth
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The tumor-forming MGC-803 cells (107 cells suspended in 100 μl PBS) transiently transfected with scramble control or siNFKB1 and siRELA were injected subcutaneously into dorsal flank of 4-week old Balb/c nude mice respectively. When the tumors were palpable in Day 4, the synthetic siRNA complex (25 nM) with siPORT Amine transfection reagent (Ambion) in 30 μl PBS was delivered intratumorally in 6-day-interval. Tumor diameter was measured and documented every 6 days until the end of Day 28. The xenografts were collected for Western blot analysis of cleaved-PARP. Tumor volume (mm3) was estimated by measuring the longest and shortest diameter of the tumor and calculating as follows: volume = (shortest diameter)2 × (longest diameter) × 0.5. All animal handling and experimental procedures were approved by Department of Health, Hong Kong (Reference No: 15–229 in DH/HA&P/8/2/1 Pt.48) and the Animal Ethics Committee of the CUHK (Reference No: 15-127-DRG).
7
siRNA Transfection and Hypoxic Conditioning of pMSCs
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The siRNA was selected in a region of the human IGF-1 transcript variants (NG-011713.1; NM-001111283 and NM-001111284). The si-IGF-1 and si-irrel RNA synthesized by Ambion (Austin, TX, USA). siRNA (100 pmol) were introduced in pMSCs that were cultured in six-well plates for 4 h by using siPORT amine transfection reagent (Ambion, Austin, TX, USA) according to the instruction of manufacturer. Forty-eight hours after transfection, the medium was changed and cells were cultured in hypoxic conditions (1% O2) for about 72 h, then the pMSCs conditoned medium for the subsequent experiments was collected.
8
Targeting miR-4534 Suppresses Prostate Cancer
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The antitumor effect of depletion of miR-4534 was determined by local administration of miR-4534 miRVANA miRNA inhibitor (antimiR) in established tumors in athymic nude mice and compared to antimiR-negative control mice group. Each mouse was injected sub-cutaneously with 2.0 ×106 Du145 prostate cancer cells. Once palpable tumors developed, 6.25 μg of synthetic anti4534 or antimiR-negative control (control) complexed with 1.6 μl siPORT Amine transfection reagent (Ambion, Austin, TX) in 50 μl PBS was delivered eight times intratumorally at 3-day intervals. In total 8 mice received anti4534 and 8 mice received control miR. Tumor growth was followed for 21 days from the first injection. All animal care was in accordance with the institutional guidelines.
9
Therapeutic Effect of miR-203 on Renal Cancer
10
Therapeutic Potential of miR-3622b in PCa
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All animal care was in accordance with the guidelines of the SFVAMC and the study was approved by the San Francisco VA IACUC. The therapeutic potential of miR-3622b was examined by local administration in established tumors in a PCa xenograft mouse model as previously described [45 (link), 47 , 48 ]. Nude mice (5 weeks-old, Simonsen Laboratories) (n=12) were injected with 2 X 106 PC3 cells (in 100 μl volume) subcutaneously in the right flanks. Once palpable tumors developed, caliper measurements were taken twice a week and tumor volumes were calculated as x2y/2, where width (x) < length (y). Synthetic miR-3622b precursor/ miR-CON (6.25 μg each) complexed with 1.6 μL siPORTamine transfection reagent (Ambion) in a volume of 50 μL PBS was delivered intratumorally every 4 days. Synthetic miRNAs are double-stranded, ready-to-use miRNA precursors and were procured from Ambion (pre-miR, cat. no. AM17100). Mice were killed 2 days after the last treatment (day 56) and tumors were harvested.
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