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Rp105

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The RP105 is a laboratory pipette used for the precise measurement and transfer of small liquid volumes. It features an ergonomic design and adjustable volume range to accommodate a variety of laboratory applications.

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Lab products found in correlation

Lipopolysaccharides (lps), by Merck Group (5 mentions) Interleukin 4 (il 4), by Merck Group (4 mentions) Sodium pyruvate, by Thermo Fisher Scientific (2 mentions) Murine il 4, by Thermo Fisher Scientific (2 mentions) Non essential amino acids (neaa), by Thermo Fisher Scientific (2 mentions) Roswell park memorial institute (rpmi), by Thermo Fisher Scientific (2 mentions) Anti cd43 microbeads, by Miltenyi Biotec (2 mentions) Penicillin streptomycin, by Thermo Fisher Scientific (2 mentions) Penicillin, by Thermo Fisher Scientific (1 mentions) Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (1 mentions) Lipopolysaccharide, by Merck Group (1 mentions) Anti b220 antibody, by BD (1 mentions) Carboxyfluorescein succinimidyl ester (cfse), by Thermo Fisher Scientific (1 mentions) Facscalibur, by BD (1 mentions) Trypan blue, by STEMCELL (1 mentions) Hemacytometer, by Hausser Scientific (1 mentions) Stem cell factor (scf), by Thermo Fisher Scientific (1 mentions) Stemspan, by Merck Group (1 mentions) β mercaptoethanol, by Thermo Fisher Scientific (1 mentions) Penicillin streptomycin, by Merck Group (1 mentions)

6 protocols using rp105

1

Culturing MEFs, HeLa, and Splenic B Cells

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MEFs and HeLa S3 cells were maintained in DMEM (Invitrogen) supplemented with 10% calf serum and 1% penicillin/streptomycin. Splenic B cells were maintained in RPMI (Invitrogen) supplemented with 10% FBS, 1% penicillin/streptomycin, 1% nonessential amino acids (Life Technologies), 1% sodium pyruvate (Life Technologies), 50 μM β-mercaptoethanol, 5 ng/ml murine IL-4 (Life Technologies), 0.5 μg/ml RP105 (552128; BD Pharmingen), and 25 μg/ml LPS (Sigma-Aldrich).
Jiang Q., Foglizzo M., Morozov Y.I., Yang X., Datta A., Tian L., Thada V., Li W., Zeqiraj E, & Greenberg R.A. (2022). Autologous K63 deubiquitylation within the BRCA1-A complex licenses DNA damage recognition. The Journal of Cell Biology, 221(9), e202111050.
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2

Murine Splenic B Cell Isolation and Retroviral Infection

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Resting splenic B cells were isolated from 8- to 12-week-old mice with anti-CD43 microbeads (anti-Ly48; Miltenyi Biotech) and were cultured with lipopolysaccharide (25 μg ml−1; Sigma) and interleukin-4 (5 ng ml−1; Sigma) and RP105 (0.5 μg ml−1; BD)41 (link). Stimulated B cells were infected with retrovirus-CRE to ensure high level of deletions of Brca2 in these cells.
Ding X., Chaudhuri A.R., Callen E., Pang Y., Biswas K., Klarmann K.D., Martin B.K., Burkett S., Cleveland L., Stauffer S., Sullivan T., Dewan A., Marks H., Tubbs A.T., Wong N., Buehler E., Akagi K., Martin S.E., Keller J.R., Nussenzweig A, & Sharan S.K. (2016). Synthetic viability by BRCA2 and PARP1/ARTD1 deficiencies. Nature Communications, 7, 12425.
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3

MEF and B cell manipulation for CSR

Cited 4 times
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Wild-type, BRCA1Δ11/Δ11, 53BP1−/− and BRCA1Δ11/Δ1153BP1−/− mouse embryonic fibroblasts (MEFs) have been described (10 (link)). Resting B cells were isolated from spleen using anti-CD43 microbeads (Miltenyi Biotec) and stimulated to proliferate with 25 μg/ml LPS, 5 ng/ml IL-4 (both Sigma-Aldrich) and 0.5 μg/ml RP105 (BD PharMingen) (10 (link)). To overexpress RNF168, constructs encoding human RNF168WT or RNF168R57D (9 (link)) were subcloned into the pMX-PIE-IRES-GFP retroviral vector and used to transfect BOSC23 cells along with the pCL-Eco helper virus. Retroviral supernatant was collected 40–48 h later for infection of MEFs and B cells as previously described (38 (link)). pMX-PIE-based retroviruses encoding 53BP1DB and 53BP1DN have been described (11 (link),28 (link)). Class switch recombination was assayed on days 3 and 4 using biotinylated anti-IgG1 and fluorochrome-conjugated anti-B220 antibodies (BD PharMingen).
Zong D., Callén E., Pegoraro G., Lukas C., Lukas J, & Nussenzweig A. (2015). Ectopic expression of RNF168 and 53BP1 increases mutagenic but not physiological non-homologous end joining. Nucleic Acids Research, 43(10), 4950-4961.
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4

Analyzing B Cell Proliferation and Genomic Instability

Cited 2 times
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B cells were isolated from WT or mutant spleen with anti-CD43 Microbeads (anti-Ly48; Miltenyi Biotec) and were cultured with LPS (25 µg/ml; Sigma-Aldrich), IL-4 (5 ng/ml; Sigma-Aldrich), and RP105 (0.5 µg/ml; BD). Cell proliferation was analyzed by CFSE (5 µM; Molecular Probes) labeling at 37°C for 10 min. For cell cycle analysis, B cells were fixed in methanol and stained with propidium iodide (PI). BrdU incorporation and detection was performed as described (Bunting et al., 2012 (link)). Samples were acquired on a FACSCalibur (BD). For genomic instability analysis, B cells were harvested after 2 d in culture; metaphase spreads were generated and processed for FISH analysis as previously described (Callen et al., 2013 (link)). PARP inhibitor (KU58948; Astra Zeneca) was added to cells stimulated ex vivo for one day.
Polato F., Callen E., Wong N., Faryabi R., Bunting S., Chen H.T., Kozak M., Kruhlak M.J., Reczek C.R., Lee W.H., Ludwig T., Baer R., Feigenbaum L., Jackson S, & Nussenzweig A. (2014). CtIP-mediated resection is essential for viability and can operate independently of BRCA1. The Journal of Experimental Medicine, 211(6), 1027-1036.
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5

Expansion and activation of hematopoietic cells

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Sorted LSK were cultured in 96-well plates containing 100 µL StemSpan supplemented with 10% FBS and 1% penicillin/streptomycin, 1% L-glutamine, 100 µM β-mercaptoethanol (Sigma), 10 ng/mL TPO, and 50 ng/mL SCF (Peprotech Inc), with or without MMC. Cells were seeded at a density of 100,000/mL in triplicate and maintained at less than 1 million/mL throughout culture by subculture into fresh media. Viable cells were counted using a Hemacytometer (Hausser Scientific) using Trypan Blue (STEMCELL technologies).
Splenic B cells were cultured in 6-well plates containing 2.5 mL RPMI (Invitrogen) supplemented with 10% FBS, 1% penicillin/streptomycin, 1% nonessential amino acids (Life Technologies), 1% sodium pyruvate (Life Technologies), 50 µM β-mercaptoethanol, 5 ng/mL murine IL-4 (Life Technologies), 0.5 µg/mL RP105 (BD Pharmingen, 552128, 1:100), and 25 µg/mL LPS (Sigma).
Balcerek J., Jiang J., Li Y., Jiang Q., Holdreith N., Singh B., Chandra V., Lv K., Ren J.G., Rozenova K., Li W., Greenberg R.A, & Tong W. (2018). Lnk/Sh2b3 deficiency restores hematopoietic stem cell function and genome integrity in Fancd2 deficient Fanconi anemia. Nature Communications, 9, 3915.
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6

Replication Dynamics in Mature B Cells

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Replication timing profiles were determined using TimEX as described (Bartholdy et al., PMID: 25987481) with minor modifications. The S/G1 TimEX ratio, proportional to the replication time during S phase, was measured in mature splenic B cells from wild-type C57BL/6NCr mice. The G1 population was obtained from mature resting B cells (100% G0/G1 arrested), which were isolated using anti-CD43 MicroBeads (Miltenyi Biotech) for negative selection. To obtain the S-phase population, the resting B cells were activated with LPS (25 mg/ml; Sigma), IL-4 (5 ng/ml; Sigma) and RP105 (0.5 mg/ml; BD) for 48 hours (~70% S-phase). 5 million cells were collected for both populations, and whole genome sequencing was performed using standard lllumina library prep and sequenced on HiSeq 2000.
Tubbs A., Sridharan S., van Wietmarschen N., Maman Y., Callen E., Stanlie A., Wu W., Wu X., Day A., Wong N., Yin M., Canela A., Fu H., Redon C., Pruitt S.C., Jaszczyszyn Y., Aladjem M.I., Aplan P.D., Hyrien O, & Nussenzweig A. (2018). Dual Roles of Poly(dA:dT) Tracts in Replication Initiation and Fork Collapse. Cell, 174(5), 1127-1142.e19.
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