RIP assays were performed to verify the targeted relationship between miR-25-3p, JPX, and PPID using the RIP Kit (Millipore Sigma) according to the manufacturer's protocol. Cells were lysed using the lysis buffer and incubated with magnetic beads pre-coated with Ago2 antibody (cat. no. ab186733; 1:1,000; Abcam) for 6 h at 4°C. IgG (cat. no. SAB5600195; 1:1,000; MilliporeSigma) was used as the control. Subsequently, beads were washed with RNA binding buffer, and the levels of JPX, miR-25-3p, and PPID were detected by RT-qPCR.
Rip kit
Manufactured by Merck Group
Sourced in United States, Germany, China
The RIP kit is a laboratory equipment product developed by Merck Group. It is designed to facilitate the analysis and purification of ribonucleoprotein (RNP) complexes. The kit provides the necessary reagents and protocols to carry out the RNA immunoprecipitation (RIP) technique, which is used to identify and study the RNA-protein interactions within a cellular environment.
Automatically generated - may contain errors
Lab products found in correlation
Ab109489, by Abcam (53 mentions)
Ab32381, by Abcam (38 mentions)
Ripa lysis buffer, by Beyotime (26 mentions)
Ab172730, by Abcam (23 mentions)
Ab186733, by Abcam (21 mentions)
Ripa lysate, by Beyotime (12 mentions)
Ab205719, by Abcam (5 mentions)
Ripa lysis, by Beyotime (4 mentions)
Trizol reagent, by Thermo Fisher Scientific (4 mentions)
Ago2 antibody, by Abcam (4 mentions)
Anti ago2, by Merck Group (3 mentions)
Ip co ip kit, by Thermo Fisher Scientific (3 mentions)
Rip lysis buffer, by Beyotime (3 mentions)
Ab13537, by Abcam (2 mentions)
Ab2410, by Abcam (2 mentions)
Anti argonaute 2 ago2 antibodies, by Abcam (2 mentions)
Trizol, by Thermo Fisher Scientific (2 mentions)
Ago2 antibody, by Merck Group (2 mentions)
Ab183403, by Abcam (2 mentions)
Human anti ago2 antibody, by Merck Group (2 mentions)
218 protocols using rip kit
1
Validating miR-25-3p, JPX, and PPID Interaction
2
Direct Interaction of LINC00630 and miR-199a
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An RIP assay was performed to explore the direct interaction between LINC00630 and miR-199a using an RIP kit (MilliporeSigma), following the manufacturer's instructions. CCA cells (2x107 cells) were lysed using RIPA lysis buffer (Beyotime Institute of Biotechnology) and then incubated with magnetic beads precoated with antibodies against Argonaute 2 (Ago2; cat. no. MABE56; Sigma-Aldrich; Merck KGaA); anti-immunoglobulin G (IgG; cat. no. I5131; Sigma-Aldrich; Merck KGaA) was used as the negative control. TRIzol reagent was used for the RNA extraction and the RNA enrichment was determined using RT-qPCR. Finally, the expression levels of LINC00630 and miR-199a in the anti-IgG and anti-Ago2 groups were compared.
3
Detection of NR2F1-AS1 Binding to SPI1
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The ability of NR2F1-AS1 to bind SPI1 was detected using the RIP kit (Millipore Sigma, Burlington, MA, USA). The cells were lysed with an equal volume of RIPA lysis (P0013B, Beyotime Biotechnology) in an ice bath for 5 min. Part of the cell extract was saved as the input, and the remaining was incubated with anti-IgG (1: 100, ab109489, Abcam) or anti-SPI1 (1: 1000, sc-365,208, Santa Cruz Biotechnology Inc.) antibodies for co-precipitation. The bead-protein complex was collected on a magnetic base. RNA was extracted from the co-precipitated samples and Input samples after detachment from the bead using protease K. The expression of NR2F1-AS1 was determined using RT-qPCR.
4
NEAT1 Binding to SPI1 Detected
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The RIP kit (Millipore Sigma, USA) was used to detect the ability of NEAT1 to bind SPI1. The hBMSCs were lysed with RIPA lysis (Beyotime Biotechnology, China) in an ice bath for 5 min. The cell extract was incubated with SPI1 (1:1000, ab227835) or IgG (1:100, ab109489) antibodies as input and the remainder was co-precipitation. A magnetic substrate was then used to collect the bead protein complex. Finally, protease K was used to detach the RNA from the co-precipitated bead protein. The expression of NEAT1, miR-339-5p and SPI1 was assessed by RT‐qPCR.
5
Validating CircRNA-miRNA Interactions
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An RIP kit (Millipore Sigma, USA) was used to validate the interaction between circ_0088212 and miR-520 h, as described in the protocol. Briefly, 80% confluent SW1353 and HOS cells were collected and treated with 200 µL of harsh lysis buffer + protease inhibitor cocktail. Cytoplasmic lysates were isolated from debris pellets and incubated with the prepared beads along with IgG or anti-AGO2 antibody at 4 °C. The beads were then collected and washed thoroughly and RT-qPCR was employed to test circ_0088212 and circ_0088212 expression.
6
RIP-based RNA Immunoprecipitation
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According to the instructions of RIP kit (Millipore, Bedford, MA, USA), Daoy and D341 cells were lysed in RIP lysate. The magnetic beads were preincubated with antibody IgG (Millipore) and anti-Ago2 (Millipore) at 37°C for 1 h. The cell lysates were immunoprecipitated by the immunoprecipitation method and overnight at 4°C, then the detection of purified immunoprecipitated RNA by qRT-PCR.
7
Characterizing lincRNA-EPS Interactions
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The RIP Kit (Millipore) was used to analyze the interaction of lincRNA-EPS with HNRNPL and MYH6 in HL-1 cells [25 (link)]. In brief, HL-1 cells were suspended in 200 μL of lysis buffer. Anti-IgG and anti-HNRNPL antibodies were used to pre-treat the magnetic beads, which were used to immunoprecipitate the cell suspension overnight at 4°C. The RNAs were purified from the protein-RNA complex and analyzed by qPCR using lincRNA-EPS and MYH6 primers.
8
Verification of H19 and miR-149 Binding to AGO2
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The binding of H19 and miR‐149 with argonaute 2 (AGO2) protein was verified according to the instructions of the RIP kit (Millipore Inc). A part of the cell extract was used as Input, and the other part was incubated with the antibodies and magnetic beads. The magnetic beads‐antibody complexes were resuspended in 900 μL RIP wash buffer. The samples were set on a magnetic base to collect the magnetic bead‐protein complexes. RNAs in the samples and input were separately extracted after detachment with protease K for the subsequent PCR detection. The antibodies for RIP included AGO2 (ab32381, 1:50, Abcam Inc) and IgG (1:100, ab109489, Abcam Inc) as the negative control.
9
LINC00184 Binds to DNMT1: RIP Assay
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The binding relationship between LINC00184 and DNMT1 was determined by RIP kit (Millipore Corp, Billerica, MA, USA) according to previous study [16 ]. The cells were lysed with RIPA lysis buffer (P0013B, Beyotime Biotechnology Co., Shanghai, China) for 5 min on ice. The lysate (100 μL) was incubated with magnetic bead coated with antibody overnight at 4 °C. Samples were placed on magnetic pedestals to collect magnetic beads-protein complexes. The immunoprecipitated bead-protein complex was digested by protease K to collect RNA, followed by RT-qPCR. The antibody used in RIP was mouse anti-DNMT1 (1:100, ab13537, Abcam) NC IgG (1:100, ab109489, Abcam).
10
BCL3-Mediated LINC00176 Enrichment
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The BCL3‐induced enrichment of LINC00176 was determined using a RIP Kit (Millipore). After a pre‐cooled PBS rinse, the HO8910 cells were lysed with equal volume of lysis in ice bath for 5 minutes. The supernatant was attained through centrifugation at 21 912 × g and 4°C for 10 minutes. A portion of the cell extract was used as input, while the remaining was probed with the BCL3 antibody (ab27780, Abcam lnc) for coprecipitation. Immunoglobulin G (IgG) antibody (ab2410, Abcam Inc) served as NC. RNA was extracted from the sample and input after protease K detachment, followed by RT‐qPCR detection.
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