We performed siRNA knockdown for HPRT1, LTB, CCL4, and KLF2 by electroporation, 6 hours prior to the MPA treatment of our lymphocyte viability assay workflow. Lymphocytes were treated with 300 nM pooled HPRT1 Stealth siRNAs (HPRT1HSS105004, HPRT1HSS105005, and HPRT1HSS105006), Silencer Select siRNAs CCL4 (s12574), KLF2 (s20269, s20270, and s20271), LTB (s8311, s8312, and s194597), or scramble control (Negative Control Median GC Duplex; Invitrogen, Carlsbad, CA) using the Amaxa P3 Primary Cell 4D‐Nucleofector X Kit S (Lonza, Walkersville, MD). The E0‐115 method was run on the 4D Nucleofector Core Unit (Lonza). Lymphocytes were maintained in T25 flasks prior to electroporation and plated into 96‐well and 6‐well plates after a 6‐hour electroporation recovery period.
Amaxa p3 primary cell 4d nucleofector x kit s
Manufactured by Lonza
The Amaxa P3 Primary Cell 4D-Nucleofector X Kit S is a laboratory equipment product designed for the transfection of primary cells. It enables the efficient delivery of nucleic acids, such as plasmid DNA or RNA, into various types of primary cells. The kit provides the necessary reagents and protocols to perform the nucleofection process.
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Lab products found in correlation
2 protocols using amaxa p3 primary cell 4d nucleofector x kit s
1
siRNA Knockdown Workflow for Lymphocyte Viability
2
Evaluating 4-1BB Knockdown in Activated PBMCs
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4-1BB siRNAs and control siRNA were synthesized by Horizon (Dharmacon Reagents, cat. nos. J-008105-08-0020 and J-008105-09-0020). Activated PBMCs (1 × 106) were transfected with 300 nM individual 4-1BB siRNA or control siRNA using an Amaxa P3 Primary Cell 4D-Nucleofector X Kit S (Lonza, cat. no. V4XP-3032) according to the manufacturer’s protocol. Transfected PBMCs were plated into a 96-well plate for 72 hours. Afterward, 105 transfected PBMCs were preincubated with 2.5 μg of human BD Fc Block (BD Biosciences, cat. no. 564220) for 10 min at RT and then stained with 1 μg of human siglec-4–Fc protein (R&D systems, cat. no. 8940-MG-050) on ice for 30 min followed by AF647 AffiniPure Donkey Anti-Human IgG (H + L) (Jackson ImmunoResearch, cat. no. 709–605-149). 4-1BB was stained using BV421 anti-CD137 (4-1BB) antibody (BioLegend, cat. no. 309819). 4-1BB expression and siglec-4 protein binding were detected using a LSRFortessa flow cytometer (BD Biosciences).
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