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Monoclonal Antibody Immunoprecipitation Mitotic Cell Cycle Checkpoint

$260
100 µl
APPLICATIONS
REACTIVITY
Human, Mouse, Rat

Application Methods: Immunoprecipitation, Western Blotting

Background: Correct segregation of sister chromatids prior to the onset of cell division is essential to the maintenance of genetic integrity and the avoidance of aneuploidy and chromosomal instability, characteristics of many cancer cells. The mitotic checkpoint, also known as the spindle assembly checkpoint, monitors accurate attachment of kinetochores to the spindle, inhibits mitosis and delays the onset of anaphase until all chromosomes are aligned at the metaphase plate (1). MAD2L1 is an essential participant in the mitotic checkpoint (2). It exists in two conformations, including the open and inactive O-MAD2 form and the closed, active C-MAD2 form. Prior to mitosis, MAD2L1 is localized to the cytosol and exists largely in the closed, inactive form. During the mitotic checkpoint, MAD2L1 switches to the open, active conformation (3). Together with other checkpoint proteins, MAD2L1 binds to and deactivates Cdc20, thereby inhibiting the anaphase promoting complex (4). When the kinetochores are correctly attached to the spindle, MAD2L1 releases Cdc20, which allows activation of the anaphase promoting complex and subsequent degradation of key mitotic substrates and the initiation of metaphase-anaphase transition (5).

$122
20 µl
$303
100 µl
APPLICATIONS
REACTIVITY
Human, Monkey, Mouse, Rat

Application Methods: Immunoprecipitation, Western Blotting

Background: The retinoblastoma tumor suppressor protein Rb regulates cell proliferation by controlling progression through the restriction point within the G1-phase of the cell cycle (1). Rb has three functionally distinct binding domains and interacts with critical regulatory proteins including the E2F family of transcription factors, c-Abl tyrosine kinase, and proteins with a conserved LXCXE motif (2-4). Cell cycle-dependent phosphorylation by a CDK inhibits Rb target binding and allows cell cycle progression (5). Rb inactivation and subsequent cell cycle progression likely requires an initial phosphorylation by cyclin D-CDK4/6 followed by cyclin E-CDK2 phosphorylation (6). Specificity of different CDK/cyclin complexes has been observed in vitro (6-8) and cyclin D1 is required for Ser780 phosphorylation in vivo (9).

$111
20 µl
$260
100 µl
$630
300 µl
APPLICATIONS
REACTIVITY
Bovine, Human, Monkey, Pig

Application Methods: Chromatin IP, Flow Cytometry, Immunofluorescence (Immunocytochemistry), Immunohistochemistry (Paraffin), Immunoprecipitation, Western Blotting

Background: The retinoblastoma tumor suppressor protein Rb regulates cell proliferation by controlling progression through the restriction point within the G1-phase of the cell cycle (1). Rb has three functionally distinct binding domains and interacts with critical regulatory proteins including the E2F family of transcription factors, c-Abl tyrosine kinase, and proteins with a conserved LXCXE motif (2-4). Cell cycle-dependent phosphorylation by a CDK inhibits Rb target binding and allows cell cycle progression (5). Rb inactivation and subsequent cell cycle progression likely requires an initial phosphorylation by cyclin D-CDK4/6 followed by cyclin E-CDK2 phosphorylation (6). Specificity of different CDK/cyclin complexes has been observed in vitro (6-8) and cyclin D1 is required for Ser780 phosphorylation in vivo (9).

$134
20 µl
$336
100 µl
APPLICATIONS
REACTIVITY
Human, Monkey, Mouse, Rat

Application Methods: Flow Cytometry, Immunofluorescence (Immunocytochemistry), Immunohistochemistry (Paraffin), Immunoprecipitation, Western Blotting

Background: The retinoblastoma tumor suppressor protein Rb regulates cell proliferation by controlling progression through the restriction point within the G1-phase of the cell cycle (1). Rb has three functionally distinct binding domains and interacts with critical regulatory proteins including the E2F family of transcription factors, c-Abl tyrosine kinase, and proteins with a conserved LXCXE motif (2-4). Cell cycle-dependent phosphorylation by a CDK inhibits Rb target binding and allows cell cycle progression (5). Rb inactivation and subsequent cell cycle progression likely requires an initial phosphorylation by cyclin D-CDK4/6 followed by cyclin E-CDK2 phosphorylation (6). Specificity of different CDK/cyclin complexes has been observed in vitro (6-8) and cyclin D1 is required for Ser780 phosphorylation in vivo (9).

$111
20 µl
$260
100 µl
APPLICATIONS
REACTIVITY
Human, Monkey, Mouse

Application Methods: Chromatin IP, Immunoprecipitation, Western Blotting

Background: The retinoblastoma tumor suppressor protein Rb regulates cell proliferation by controlling progression through the restriction point within the G1-phase of the cell cycle (1). Rb has three functionally distinct binding domains and interacts with critical regulatory proteins including the E2F family of transcription factors, c-Abl tyrosine kinase, and proteins with a conserved LXCXE motif (2-4). Cell cycle-dependent phosphorylation by a CDK inhibits Rb target binding and allows cell cycle progression (5). Rb inactivation and subsequent cell cycle progression likely requires an initial phosphorylation by cyclin D-CDK4/6 followed by cyclin E-CDK2 phosphorylation (6). Specificity of different CDK/cyclin complexes has been observed in vitro (6-8) and cyclin D1 is required for Ser780 phosphorylation in vivo (9).

$303
100 µl
APPLICATIONS
REACTIVITY
Human, Monkey, Mouse, Rat

Application Methods: Immunoprecipitation, Western Blotting

Background: The retinoblastoma tumor suppressor protein Rb regulates cell proliferation by controlling progression through the restriction point within the G1-phase of the cell cycle (1). Rb has three functionally distinct binding domains and interacts with critical regulatory proteins including the E2F family of transcription factors, c-Abl tyrosine kinase, and proteins with a conserved LXCXE motif (2-4). Cell cycle-dependent phosphorylation by a CDK inhibits Rb target binding and allows cell cycle progression (5). Rb inactivation and subsequent cell cycle progression likely requires an initial phosphorylation by cyclin D-CDK4/6 followed by cyclin E-CDK2 phosphorylation (6). Specificity of different CDK/cyclin complexes has been observed in vitro (6-8) and cyclin D1 is required for Ser780 phosphorylation in vivo (9).

$122
20 µl
$303
100 µl
APPLICATIONS
REACTIVITY
Human

Application Methods: Immunoprecipitation, Western Blotting

Background: The retinoblastoma tumor suppressor protein Rb regulates cell proliferation by controlling progression through the restriction point within the G1-phase of the cell cycle (1). Rb has three functionally distinct binding domains and interacts with critical regulatory proteins including the E2F family of transcription factors, c-Abl tyrosine kinase, and proteins with a conserved LXCXE motif (2-4). Cell cycle-dependent phosphorylation by a CDK inhibits Rb target binding and allows cell cycle progression (5). Rb inactivation and subsequent cell cycle progression likely requires an initial phosphorylation by cyclin D-CDK4/6 followed by cyclin E-CDK2 phosphorylation (6). Specificity of different CDK/cyclin complexes has been observed in vitro (6-8) and cyclin D1 is required for Ser780 phosphorylation in vivo (9).

$260
100 µl
APPLICATIONS
REACTIVITY
Human

Application Methods: Immunoprecipitation, Western Blotting

Background: At least five distinct polo-like kinases exist in mammalian cells: PLK1, PLK2, PLK3, PLK4/SAK, and the non-catalytic PLK5 protein (1). The p53-induced PLK2 functions in centriole duplication, as well as at spindle and S phase checkpoints (3-5). Research studies show that PLK2 expression is related to chemosensitivity in ovarian cancer. Downregulation of PLK2 expression in chemosensitive ovarian cancer cells is associated with a greater chance of relapse in patients following specific treatment regimens (6). PLK2 can phosphorylate α-synuclein at Ser129, which is a site shown to be involved in diseases of the central nervous system (7,8). Polo-like kinase 2 also phosphorylates GEFs and GAPs, regulating Ras and Rap small GTPase function in neurons (9).

$260
100 µl
APPLICATIONS
REACTIVITY
Human, Mouse

Application Methods: Immunoprecipitation, Western Blotting

Background: At least 4 distinct polo-like kinases exist in mammalian cells: PLK1, PLK2, PLK3 and PLK4/SAK (1). Like the other PLK family members, PLK3 contains an amino-terminal catalytic domain and a conserved carboxy-terminal domain termed the Polo box (2). PLK3, also called proliferation-related kinase (Prk) (3), was originally described as a fibroblast growth factor (FGF)-inducible kinase (Fnk) and identified as an immediate-early response gene responsive to FGF-1 and other mitogens (4). PLK3 is a cytokine-inducible serine/threonine kinase whose protein expression is cell cycle regulated. Though its expression is found primarily in G1 phase of the cell cycle, PLK3 is detected in G0 and in late telophase prior to cytokinesis (5). Like the other PLK family members, PLK3 functions mainly as a regulator of the cell cycle. Specifically, PLK3 is required for entry into S phase and is a critical regulator of G1 events, as indicated by RNAi-induced PLK3-depleted cells (2). PLK3 is also involved in the regulation of DNA damage response via phosphorylation of p53 on Ser20 (6). PLK3 may act as a tumor suppressor as Plk3-deficient mice develop spontaneous tumors in various organs (7). Unlike PLK1, PLK3 expression is down regulated in cancers including lung (3), head and neck (8), and colon (9).

$260
100 µl
APPLICATIONS
REACTIVITY
Human, Mouse, Rat

Application Methods: Immunofluorescence (Immunocytochemistry), Immunoprecipitation, Western Blotting

Background: Nijmegen breakage syndrome (NBS) is characterized by growth retardation, mental disability, immunodeficiency, defects in cell cycle checkpoints, an increased propensity for cancer, and sensitivity to ionizing radiation (1). Repair of radiation-induced DNA double-strand breaks is dependent on the multifunctional MRN complex containing Mre11, Rad50, and the NBS1 gene product p95/NBS1 (also called p95 or nibrin) (2). p95/NBS1 is a protein with a forkhead-associated domain and a BRCT repeat that regulate interaction with MDC1 and are essential for proper G2/M DNA-damage checkpoint function (3). NBS1 is critical for homologous recombination following DNA double strand breaks. This activity requires CDK-dependent association with CtIP and subsequent phosphorylation by ATM (4). ATM interacts with and phosphorylates p95/NBS1 at Ser278 and Ser343 after exposure to ionizing radiation (5,6).

$260
100 µl
APPLICATIONS
REACTIVITY
Human, Monkey, Mouse, Rat

Application Methods: Immunoprecipitation, Western Blotting

Background: Checkpoint with forkhead and RING finger domains protein (CHFR) is an E3 ubiquitin-protein ligase that regulates cell cycle progression. In response to microtubule stress, CHFR delays the transition into mitosis by excluding cyclin B1 from the nucleus prior to chromosome condensation (1). Marked reduction of CHFR expression was detected in primary tumors and decreased CHFR expression was linked to promoter hypermethylation (1-4). Restoration of CHFR expression by treatment with the microtubule stress agent nocodazole and the methyltransferase inhibitor 5-aza-2'-deoxycytidine has been reported (4,5).

$303
100 µl
APPLICATIONS
REACTIVITY
Human

Application Methods: Immunoprecipitation, Western Blotting

Background: The human checkpoint protein Rad17 and its fission and budding yeast orthologues (Schizosaccharomyces pombe Rad17 and Saccharomyces cerevisiae Rad24, respectively) are involved in the activation of checkpoint signals in response to DNA damage or disruption of DNA synthesis (1-4). Treatment of human cells with genotoxic agents induces ATM/ATR-dependent phosphorylation of Rad17 at Ser635 and Ser645. Rad17 phosphorylation is a critical early event during checkpoint signaling in DNA-damaged cells (5-7).

$111
20 µl
$260
100 µl
APPLICATIONS
REACTIVITY
Human, Monkey, Mouse, Rat

Application Methods: Flow Cytometry, Immunofluorescence (Immunocytochemistry), Immunohistochemistry (Paraffin), Immunoprecipitation, Western Blotting

Background: Bcl-xL prevents apoptosis through two different mechanisms: heterodimerization with an apoptotic protein inhibits its apoptotic effect (1,2) and formation of mitochondrial outer membrane pores help maintain a normal membrane state under stressful conditions (3). Bcl-xL is phosphorylated by JNK following treatment with microtubule-damaging agents such as paclitaxel, vinblastine and nocodazole (4,5).

$303
100 µl
APPLICATIONS
REACTIVITY
Human

Application Methods: Immunoprecipitation, Western Blotting

Background: At least four distinct polo-like kinases exist in mammalian cells: PLK1, PLK2, PLK3, and PLK4/SAK (1). PLK1 apparently plays many roles during mitosis, particularly in regulating mitotic entry and exit. The mitosis promoting factor (MPF), cdc2/cyclin B1, is activated by dephosphorylation of cdc2 (Thr14/Tyr15) by cdc25C. PLK1 phosphorylates cdc25C at Ser198 and cyclin B1 at Ser133 causing translocation of these proteins from the cytoplasm to the nucleus (2-5). PLK1 phosphorylation of Myt1 at Ser426 and Thr495 has been proposed to inactivate Myt1, one of the kinases known to phosphorylate cdc2 at Thr14/Tyr15 (6). Polo-like kinases also phosphorylate the cohesin subunit SCC1, causing cohesin displacement from chromosome arms that allow for proper cohesin localization to centromeres (7). Mitotic exit requires activation of the anaphase promoting complex (APC) (8), a ubiquitin ligase responsible for removal of cohesin at centromeres, and degradation of securin, cyclin A, cyclin B1, Aurora A, and cdc20 (9). PLK1 phosphorylation of the APC subunits Apc1, cdc16, and cdc27 has been demonstrated in vitro and has been proposed as a mechanism by which mitotic exit is regulated (10,11).Substitution of Thr210 with Asp has been reported to elevate PLK1 kinase activity and delay/arrest cells in mitosis, while a Ser137Asp substitution leads to S-phase arrest (12). In addition, while DNA damage has been found to inhibit PLK1 kinase activity, the Thr210Asp mutant is resistant to this inhibition (13). PLK1 has been reported to be phosphorylated in vivo at Ser137 and Thr210 in mitosis; DNA damage prevents phosphorylation at these sites (14).

$260
100 µl
APPLICATIONS
REACTIVITY
Human, Monkey, Mouse, Rat

Application Methods: Immunoprecipitation, Western Blotting

Background: Eukaryotic cell proliferation depends strictly upon the E3 ubiquitin ligase activity of the anaphase promoting complex/cyclosome (APC/C), whose main function is to trigger the transition of the cell cycle from metaphase to anaphase. The APC/C complex promotes the assembly of polyubiquitin chains on substrate proteins in order to target these proteins for degradation by the 26S proteasome (1,2). The vertebrate APC/C complex consists of as many as 15 subunits, including multiple scaffold proteins, two catalytic subunits (APC2, APC11), and a number of proteins responsible for substrate recognition (3). All E3 enzymes, including APC/C, utilize ubiquitin residues activated by E1 enzymes and transferred to E2 enzymes. Research studies indicate that APC/C interacts with the E2 enzymes UBE2S and UBE2C via the RING-finger domain-containing subunit APC11 (4-6). APC/C function relies on multiple cofactors, including an APC/C coactivator formed by the cell division control protein 20 homolog (CDC20) and Cdh1/FZR1. The CDC20/Cdh1 coactivator is responsible for recognition of APC/C substrates through interaction with specific D-box and KEN-box recognition elements within these substrates (7-9).

$260
100 µl
APPLICATIONS
REACTIVITY
Hamster, Human, Monkey, Mouse, Rat

Application Methods: Immunoprecipitation, Western Blotting

Background: The entry of eukaryotic cells into mitosis is regulated by cdc2 kinase activation, a process controlled at several steps including cyclin binding and phosphorylation of cdc2 at Thr161 (1). However, the critical regulatory step in activating cdc2 during progression into mitosis appears to be dephosphorylation of cdc2 at Thr14 and Tyr15 (2). Phosphorylation at Thr14 and Tyr15, resulting in inhibition of cdc2, can be carried out by Wee1 and Myt1 protein kinases (3,4). The cdc25 phosphatase may be responsible for removal of phosphates at Thr14 and Tyr15 and subsequent activation of cdc2 (1,5).

$260
100 µl
APPLICATIONS
REACTIVITY
Human, Monkey, Mouse, Rat

Application Methods: Immunoprecipitation, Western Blotting

Background: The cell division cycle demands accuracy to avoid the accumulation of genetic damage. This process is controlled by molecular circuits called "checkpoints" that are common to all eukaryotic cells (1). Checkpoints monitor DNA integrity and cell growth prior to replication and division at the G1/S and G2/M transitions, respectively. The cdc2-cyclin B kinase is pivotal in regulating the G2/M transition (2,3). Cdc2 is phosphorylated at Thr14 and Tyr15 during G2-phase by the kinases Wee1 and Myt1, rendering it inactive. The tumor suppressor protein retinoblastoma (Rb) controls progression through the late G1 restriction point (R) and is a major regulator of the G1/S transition (4). During early and mid G1-phase, Rb binds to and represses the transcription factor E2F (5). The phosphorylation of Rb late in G1-phase by CDKs induces Rb to dissociate from E2F, permitting the transcription of S-phase-promoting genes. In vitro, Rb can be phosphorylated at multiple sites by cdc2, cdk2, and cdk4/6 (6-8). DNA damage triggers both the G2/M and the G1/S checkpoints. DNA damage activates the DNA-PK/ATM/ATR kinases, which phosphorylate Chk at Ser345 (9), Chk2 at Thr68 (10) and p53 (11). The Chk kinases inactivate cdc25 via phosphorylation at Ser216, blocking the activation of cdc2.

$260
100 µl
APPLICATIONS
REACTIVITY
Human, Monkey

Application Methods: Immunoprecipitation, Western Blotting

Background: Eukaryotic cell proliferation depends strictly upon the E3 ubiquitin ligase activity of the anaphase promoting complex/cyclosome (APC/C), whose main function is to trigger the transition of the cell cycle from metaphase to anaphase. The APC/C complex promotes the assembly of polyubiquitin chains on substrate proteins in order to target these proteins for degradation by the 26S proteasome (1,2). The vertebrate APC/C complex consists of as many as 15 subunits, including multiple scaffold proteins, two catalytic subunits (APC2, APC11), and a number of proteins responsible for substrate recognition (3). All E3 enzymes, including APC/C, utilize ubiquitin residues activated by E1 enzymes and transferred to E2 enzymes. Research studies indicate that APC/C interacts with the E2 enzymes UBE2S and UBE2C via the RING-finger domain-containing subunit APC11 (4-6). APC/C function relies on multiple cofactors, including an APC/C coactivator formed by the cell division control protein 20 homolog (CDC20) and Cdh1/FZR1. The CDC20/Cdh1 coactivator is responsible for recognition of APC/C substrates through interaction with specific D-box and KEN-box recognition elements within these substrates (7-9).

$111
20 µl
$260
100 µl
APPLICATIONS
REACTIVITY
Human, Monkey, Rat

Application Methods: Immunohistochemistry (Paraffin), Immunoprecipitation, Western Blotting

Background: At least four distinct polo-like kinases exist in mammalian cells: PLK1, PLK2, PLK3, and PLK4/SAK (1). PLK1 apparently plays many roles during mitosis, particularly in regulating mitotic entry and exit. The mitosis promoting factor (MPF), cdc2/cyclin B1, is activated by dephosphorylation of cdc2 (Thr14/Tyr15) by cdc25C. PLK1 phosphorylates cdc25C at Ser198 and cyclin B1 at Ser133 causing translocation of these proteins from the cytoplasm to the nucleus (2-5). PLK1 phosphorylation of Myt1 at Ser426 and Thr495 has been proposed to inactivate Myt1, one of the kinases known to phosphorylate cdc2 at Thr14/Tyr15 (6). Polo-like kinases also phosphorylate the cohesin subunit SCC1, causing cohesin displacement from chromosome arms that allow for proper cohesin localization to centromeres (7). Mitotic exit requires activation of the anaphase promoting complex (APC) (8), a ubiquitin ligase responsible for removal of cohesin at centromeres, and degradation of securin, cyclin A, cyclin B1, Aurora A, and cdc20 (9). PLK1 phosphorylation of the APC subunits Apc1, cdc16, and cdc27 has been demonstrated in vitro and has been proposed as a mechanism by which mitotic exit is regulated (10,11).Substitution of Thr210 with Asp has been reported to elevate PLK1 kinase activity and delay/arrest cells in mitosis, while a Ser137Asp substitution leads to S-phase arrest (12). In addition, while DNA damage has been found to inhibit PLK1 kinase activity, the Thr210Asp mutant is resistant to this inhibition (13). PLK1 has been reported to be phosphorylated in vivo at Ser137 and Thr210 in mitosis; DNA damage prevents phosphorylation at these sites (14).

$122
20 µl
$303
100 µl
$717
300 µl
APPLICATIONS
REACTIVITY
Human, Monkey, Mouse, Rat

Application Methods: Flow Cytometry, Immunofluorescence (Immunocytochemistry), Immunoprecipitation, Western Blotting

Background: The entry of eukaryotic cells into mitosis is regulated by cdc2 kinase activation, a process controlled at several steps including cyclin binding and phosphorylation of cdc2 at Thr161 (1). However, the critical regulatory step in activating cdc2 during progression into mitosis appears to be dephosphorylation of cdc2 at Thr14 and Tyr15 (2). Phosphorylation at Thr14 and Tyr15, resulting in inhibition of cdc2, can be carried out by Wee1 and Myt1 protein kinases (3,4). The cdc25 phosphatase may be responsible for removal of phosphates at Thr14 and Tyr15 and subsequent activation of cdc2 (1,5).