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Normal mouse igg1

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Normal mouse IgG1 is a laboratory reagent used as a control for immunoassays and other experimental techniques. It is a purified immunoglobulin G1 (IgG1) isotype derived from normal mouse serum.

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Lab products found in correlation

Halt protease inhibitor, by Thermo Fisher Scientific (3 mentions) Phenyl methyl sulfonyl fluoride (pmsf), by Merck Group (3 mentions) Anti cdk2, by Santa Cruz Biotechnology (3 mentions) Anti prb s807 s811 antibody, by Cell Signaling Technology (3 mentions) Protease inhibitor, by Thermo Fisher Scientific (1 mentions) Dynabeads protein g, by Thermo Fisher Scientific (1 mentions) Dynamag 2 magnet, by Thermo Fisher Scientific (1 mentions)

Close spelling variants of this product

Mouse IgG1 (9 citations)

4 protocols using normal mouse igg1

1

Quantifying CDK2 Kinase Activity in PDAC Cells

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To analyze the CDK2 kinase activity, primary PDAC cells that were exposed to DMSO or palbociclib (200 nM) for 48 hour were lysed using the kinase lysis buffer (50 mM HEPES-KOH pH 7.5, 150 mM NaCl, 1 mM EDTA, 1mM DTT, 0.1% Tween-20) in the presence of 1X Halt protease inhibitor (Thermo Fisher) and 1 mM PMSF (Sigma). Active CDK2 complex was immunoprecipitated by incubating 300 μg of the lysate with 5 μg of anti-CDK2 (Santa Cruz Biotechnology; SC-6248) overnight at 4°C. Normal mouse IgG1 (Cell Signaling Technology, 5415) was used as a control. Protein G agarose-beads were added to each IP samples and incubated up to 4 hour at 4°C. Protein immunocomplexes were washed 3 times with the kinase lysis buffer and 2 times with kinase reaction buffer (40 mM Tris-HCl pH 8, 20 mM MgCl2, 0.1 mg/mL BSA, 50 μM DTT). Kinase reactions were carried out in 100 μl of kinase buffer in the presence of 100 μM ATP and 0.5 μg of bacterially purified RB C-terminal as substrate25 (link) by gently shaking at room temperature up to 30 minutes. The resulting phosphorylated RB protein was detected by immunoblotting using anti-pRb (S807/S811) antibody (Cell Signaling Technology, 8516S).
Knudsen E.S., Kumarasamy V., Ruiz A., Sivinski J., Chung S., Grant A., Vail P., Chauhan S.S., Jie T., Riall T.S, & Witkiewicz A.K. (2019). Cell cycle plasticity driven by MTOR signaling: Integral resistance to CDK4/6 inhibition in patient-derived models of pancreatic cancer. Oncogene, 38(18), 3355-3370.
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2

Quantifying CDK2 Kinase Activity in PDAC Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols
To analyze the CDK2 kinase activity, primary PDAC cells that were exposed to DMSO or palbociclib (200 nM) for 48 hour were lysed using the kinase lysis buffer (50 mM HEPES-KOH pH 7.5, 150 mM NaCl, 1 mM EDTA, 1mM DTT, 0.1% Tween-20) in the presence of 1X Halt protease inhibitor (Thermo Fisher) and 1 mM PMSF (Sigma). Active CDK2 complex was immunoprecipitated by incubating 300 μg of the lysate with 5 μg of anti-CDK2 (Santa Cruz Biotechnology; SC-6248) overnight at 4°C. Normal mouse IgG1 (Cell Signaling Technology, 5415) was used as a control. Protein G agarose-beads were added to each IP samples and incubated up to 4 hour at 4°C. Protein immunocomplexes were washed 3 times with the kinase lysis buffer and 2 times with kinase reaction buffer (40 mM Tris-HCl pH 8, 20 mM MgCl2, 0.1 mg/mL BSA, 50 μM DTT). Kinase reactions were carried out in 100 μl of kinase buffer in the presence of 100 μM ATP and 0.5 μg of bacterially purified RB C-terminal as substrate25 (link) by gently shaking at room temperature up to 30 minutes. The resulting phosphorylated RB protein was detected by immunoblotting using anti-pRb (S807/S811) antibody (Cell Signaling Technology, 8516S).
Knudsen E.S., Kumarasamy V., Ruiz A., Sivinski J., Chung S., Grant A., Vail P., Chauhan S.S., Jie T., Riall T.S, & Witkiewicz A.K. (2019). Cell cycle plasticity driven by MTOR signaling: Integral resistance to CDK4/6 inhibition in patient-derived models of pancreatic cancer. Oncogene, 38(18), 3355-3370.
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3

Immunoprecipitation for α-Synuclein in KD Patients

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Human tissue (2 infantile KD patients, 1 late-onset KD patient, and an infantile control) was obtained from the University of Maryland Brain Tissue Bank. Immunoprecipitation was performed using Dynabeads protein G (cat# 10007D, ThermoFisher Scientific) and a Dynamag-2 magnet (cat# 12321D, ThermoFisher Scientific). Initially, 50 µl of protein G beads were washed and incubated overnight at 4 °C with 5 µg of anti-α-synuclein (cat# MA1-90342, ThermoFisher Scientific) or normal mouse IgG1 (cat# 5415, Cell Signaling). Beads were washed and then incubated overnight at 4 °C with tissue lysate. Lysate was prepared from about 250 mg tissue from each sample homogenized in 30 volumes of sterile PBS with protease inhibitors (cat# 88668, ThermoFisher Scientific) using a T8.10 disperser polytron tissue homogenizer (IKA Works, Inc., Wilmington, NC). After incubation, lysate was removed and beads were washed 3 times with 200 μl PBS and transferred to a clean tube. PBS buffer was then removed and beads were used for psychosine extraction or protein elution.
Abdelkarim H., Marshall M.S., Scesa G., Smith R.A., Rue E., Marshall J., Elackattu V., Stoskute M., Issa Y., Santos M., Nguyen D., Hauck Z., van Breemen R., Celej M.S., Gaponenko V, & Bongarzone E.R. (2018). α-Synuclein interacts directly but reversibly with psychosine: implications for α-synucleinopathies. Scientific Reports, 8, 12462.
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4

Kinase Activity Assay for CDK2 in PDAC Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols
To analyse the CDK2 kinase activity, primary PDAC cells were lysed using
the kinase lysis buffer (50 mM HEPES-KOH pH 7.5, 150 mM NaCl, 1 mM EDTA, 1 mM
DTT, 0.1% Tween-20) in the presence of 1X Halt protease inhibitor (Thermo
Fisher) and 1 mM PMSF (Sigma). Active CDK2 complex was immunoprecipitated by
incubating 300 μg of the lysate with 5 μg of anti-CDK2 (Santa Cruz
Biotechnology, SC-6248) overnight at 4°C. Normal mouse IgG1 (Cell
Signaling Technology, 5415) was used as a control. Protein G agarose-beads were
added to each IP samples and incubated up to 4 hour at 4°C. Protein
immuno-complexes were washed three times with the kinase lysis buffer and two
times with kinase reaction buffer (40 mM Tris-HCl pH 8, 20 mM MgCl2,
0.1 mg/mL BSA, 50 μM DTT). Kinase reactions were carried out in 100
μL of kinase buffer in the presence of 100 μM ATP and 0.5
μg of RB protein as substrate by gently shaking at room temperature up to
30 min. The resulting phosphorylated RB protein was detected by immunoblotting
using anti-p-Rb (S807/S811) antibody (Cell Signaling Technology, 8516S).
Knudsen E.S., Kumarasamy V., Chung S., Ruiz A., Vail P., Tzetzo S., Wu J., Nambiar R., Seshadri M., Abrams S.I., Wang J, & Witkiewicz A.K. (2020). Targeting dual signalling pathways in concert with immune checkpoints for the treatment of pancreatic cancer. Gut, 70(1), 127-138.
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