KIF2C Antibody

What is KIF2C and why is it important in research?

KIF2C (Kinesin Family Member 2C) is a protein that regulates microtubule dynamics during cell division, which is essential for accurate chromosome segregation. It associates with the centromere during early prophase and disassociates after telophase, ensuring proper chromosome alignment and separation . KIF2C is particularly abundant in thymus and testis tissues, with lower expression in small intestine, colon mucosal lining, and placenta . Its interaction with microtubules and other proteins involved in mitotic spindle assembly makes it vital for maintaining genomic stability . Dysregulation of KIF2C has been implicated in various cancers, suggesting its potential as a therapeutic target . The gene for human KIF2C is mapped to chromosome 1p34.1, highlighting its genetic significance in research contexts .

What applications are KIF2C antibodies validated for?

KIF2C antibodies have been validated for multiple laboratory applications based on the search results. The polyclonal antibody (28372-1-AP) is validated for Western Blot (WB), Immunohistochemistry (IHC), and ELISA applications . It shows reactivity with human, mouse, and rat samples . For the phospho-specific antibody targeting Ser95, validated applications include Western Blot (WB), Immunocytochemistry/Immunofluorescence (ICC/IF), and ELISA, with reactivity for human and mouse samples . The monoclonal antibody (2488C3a) has been validated for Western Blot (WB), Immunoprecipitation (IP), Immunofluorescence (IF), and Flow Cytometry (FCM), specifically detecting KIF2C protein of human origin .

What are the recommended dilutions for different applications?

The recommended dilutions vary depending on the specific KIF2C antibody and application:

It is recommended that these reagents should be titrated in each testing system to obtain optimal results, as results may be sample-dependent .

What are the proper storage conditions for KIF2C antibodies?

For optimal preservation of antibody activity, proper storage is essential. The polyclonal antibody (28372-1-AP) should be stored at -20°C and is stable for one year after shipment. The manufacturer notes that aliquoting is unnecessary for -20°C storage, and the 20μl size contains 0.1% BSA . The antibody is supplied in PBS with 0.02% sodium azide and 50% glycerol at pH 7.3 . For the phospho-specific antibody (A94243), it should be shipped at 4°C, and upon delivery, it should be aliquoted and stored at -20°C. Freeze/thaw cycles should be avoided . This antibody is supplied in Phosphate Buffered Saline (without Mg²⁺ and Ca²⁺), pH 7.4, with 150mM NaCl, 0.02% Sodium Azide, and 50% Glycerol .

How does KIF2C expression correlate with cancer prognosis?

Research has revealed significant statistical correlations between KIF2C expression and clinical prognosis in various cancers . Analysis using multiple databases including TIMER, TCGA, and GEPIA2 demonstrated that KIF2C is frequently abnormally expressed in numerous cancer types . KIF2C expression is upregulated in many tumor tissues compared to corresponding normal tissues, including BLCA, BRCA, CESC, CHOL, COAD, DLBC, ESCA, GBM, HNSC, KIRC, KIRP, LIHC, LUAD, LUSC, OV, PAAD, PCPG, PRAD, READ, SARC, SKCM, STAD, THYM, UCEC, and UCS . Conversely, KIF2C expression is downregulated in some tumor tissues such as TCGT and LAML .

What are the genetic alteration patterns of KIF2C in different cancers?

The genetic alteration patterns of KIF2C vary across different cancer types, with implications for research design and interpretation. According to analysis from the cBioPortal database, the highest mutation frequency (>6%) was observed in uterine corpus endometrial carcinoma (UCEC) . The predominant mutation type in pancreatic-gastric-pedunculated carcinoma (PGPC) was a deep deletion resulting in a frameshift .

Amplification was the major type of copy number alteration (CNA) in ovarian cancer (OV), showing an alteration frequency of approximately 4% . All cases of colon adenocarcinoma (COAD) and acute myeloid leukemia (LAML) with genetic alterations had mutations in KIF2C . Additionally, amplification of KIF2C was present in all cases of sarcoma (SARC), mesothelioma (MESO), and liver hepatocellular carcinoma (LIHC) .

The missense mutation was identified as the only type of genetic alteration of KIF2C, suggesting a specific mechanism of functional change . These genetic alteration patterns provide important insights for researchers designing studies on KIF2C in different cancer contexts.

What protocols are recommended for using KIF2C antibodies in IHC applications?

For immunohistochemistry applications using KIF2C antibodies, specific protocols have been validated and published. In one published study, tissue sections were incubated with antibody against KIF2C (28372-1-AP; Proteintech) at a 1:200 dilution for 1 hour at room temperature . Following primary antibody incubation, sections were treated with MaxVision HRP-Polymer anti-Rabbit IHC Kit (KIT-5030, MXB Biotechnologies) for 30 minutes . For visualization, the working solution of DAB (DAB-2032, MXB Biotechnologies) was applied to the tissue sections for the chromogenic reaction . The tissue sections were then examined using an upright microscope (BX53, Olympus) .

For optimal antigen retrieval, it is suggested to use TE buffer at pH 9.0, though citrate buffer at pH 6.0 may alternatively be used . The recommended dilution range for IHC applications is 1:50-1:500, but researchers should titrate the antibody in their specific testing systems to achieve optimal results . Positive IHC detection has been confirmed in mouse testis tissue .

How do phospho-specific KIF2C antibodies differ from pan-KIF2C antibodies in research applications?

Phospho-specific KIF2C antibodies target KIF2C only when phosphorylated at specific residues, enabling researchers to study post-translational modifications that may affect protein function. The phospho-Ser95 antibody (A94243) specifically detects endogenous levels of KIF2C only when phosphorylated at Ser95 . This antibody was generated using a synthetic peptide derived from human KIF2C around the phosphorylation site of Ser95 (amino acids 61-110) .

In contrast, pan-KIF2C antibodies such as 28372-1-AP detect the KIF2C protein regardless of its phosphorylation status . This fundamental difference allows researchers to design experiments that can distinguish between total KIF2C protein levels and the proportion that is phosphorylated at specific residues, providing insights into signaling pathways and regulatory mechanisms that control KIF2C function.

The phospho-specific antibody has been purified from rabbit serum by antigen affinity chromatography using the immunizing phospho peptide , ensuring specificity for the phosphorylated form. This specificity makes phospho-KIF2C antibodies valuable tools for investigating the role of phosphorylation in regulating KIF2C's activity during mitosis and in cancer progression.

What are common issues when using KIF2C antibodies and how can they be resolved?

When working with KIF2C antibodies, researchers may encounter several common issues that can affect experimental outcomes. One frequent challenge is background staining or non-specific binding in immunohistochemistry and immunofluorescence applications. To minimize this, optimizing blocking conditions is essential—typically using 5-10% normal serum from the same species as the secondary antibody for 1-2 hours at room temperature.

Antibody specificity can also be a concern, particularly when studying closely related kinesin family members. Validation using positive and negative controls is crucial, with mouse testis tissue serving as a reliable positive control for KIF2C antibodies . Additionally, verifying specificity through knockdown/knockout experiments can provide confidence in antibody performance, as several publications have used KIF2C antibodies in such applications .

For western blots, the observed molecular weight of KIF2C is approximately 80 kDa , which aligns with the calculated molecular weight of 81 kDa (725 amino acids) . If bands at unexpected molecular weights appear, they may represent degradation products, post-translationally modified forms, or non-specific binding. Optimizing primary antibody concentration, incubation time, and washing steps can help resolve these issues.

How can researchers validate the specificity of KIF2C antibodies?

Validating antibody specificity is critical for reliable research outcomes. For KIF2C antibodies, researchers should implement multiple validation strategies. First, western blot analysis using positive control samples such as DU 145 cells, PC-3 cells, mouse testis, or rat testis can confirm detection of the expected 80 kDa band .

Second, knockout/knockdown validation provides robust confirmation of antibody specificity. Several publications have used KIF2C antibodies in KD/KO applications , suggesting these antibodies can reliably distinguish between samples with and without KIF2C expression.

Third, immunohistochemistry should show appropriate subcellular localization patterns. KIF2C is primarily located in both the cellular nucleus and cytosol , and during mitosis, it associates with the centromere during early prophase and disassociates after telophase . Proper localization can be verified using confocal microscopy with appropriate co-staining markers.

For phospho-specific antibodies, validation should include treatment with phosphatase to confirm that signal loss occurs when phosphorylation is removed. Additionally, stimulating cells with agents known to induce the specific phosphorylation event can demonstrate antibody responsiveness to physiological changes in phosphorylation status.

What are the best practices for multiplexing KIF2C antibodies with other markers?

When designing multiplex experiments using KIF2C antibodies alongside other markers, several considerations can optimize results. First, antibody compatibility must be ensured—choose primary antibodies raised in different host species to prevent cross-reactivity of secondary antibodies. Based on the search results, KIF2C antibodies are available as rabbit polyclonal (28372-1-AP) , rabbit polyclonal phospho-specific (A94243) , and mouse monoclonal (2488C3a) , providing flexibility in experimental design.

Sequential staining protocols may be necessary when using multiple rabbit antibodies. This involves complete staining with the first primary and secondary antibody pair, followed by elution or inactivation of these antibodies before applying the second set. Tyramide signal amplification can be useful in these scenarios, as it allows permanent deposition of fluorophores that remain after antibody elution.

For optimal multiplexing, consider spectral characteristics of fluorophores when using immunofluorescence—choose fluorophores with minimal spectral overlap and include proper controls for autofluorescence and bleed-through. When working with tissue samples, be aware that KIF2C shows tissue-specific expression patterns, being particularly abundant in thymus and testis while showing lower expression in small intestine, colon, and placenta .

How is KIF2C being studied as a potential therapeutic target in cancer?

KIF2C has emerged as a promising therapeutic target in cancer research due to its critical role in cell division and its aberrant expression in various malignancies. Multi-omics analysis has demonstrated significant statistical correlations between KIF2C expression and clinical prognosis in multiple cancer types . The elevated expression of KIF2C in numerous tumors compared to corresponding normal tissues suggests its potential oncogenic role .

Researchers are exploring several approaches to target KIF2C therapeutically. One strategy involves developing small molecule inhibitors that can disrupt KIF2C's microtubule-depolymerizing activity, potentially halting mitosis in rapidly dividing cancer cells. Another approach focuses on degrading KIF2C protein using proteolysis-targeting chimeras (PROTACs) or related technologies.

What role does KIF2C phosphorylation play in cellular function?

Phosphorylation of KIF2C plays a crucial role in regulating its activity and function during cell division. The availability of phospho-specific antibodies targeting Ser95 suggests the importance of this particular phosphorylation site in KIF2C regulation . Phosphorylation can modulate KIF2C's microtubule-depolymerizing activity, which is essential for proper chromosome alignment and segregation during mitosis.

Research using phospho-specific antibodies has enabled investigators to study how signaling pathways regulate KIF2C through post-translational modifications. These studies provide insights into how KIF2C activity is precisely controlled during the cell cycle and how dysregulation of these phosphorylation events might contribute to genomic instability and cancer development.

The phospho-Ser95 antibody specifically detects endogenous levels of KIF2C only when phosphorylated at Ser95 , allowing researchers to track this specific modification in various experimental contexts. This site-specific phosphorylation may affect KIF2C's interactions with binding partners, its subcellular localization, or its enzymatic activity. Understanding these regulatory mechanisms could reveal new approaches for therapeutic intervention in diseases characterized by aberrant KIF2C function.

How can multi-omics approaches advance KIF2C research?

Multi-omics approaches have already shown significant value in KIF2C research, as demonstrated by studies that have performed comprehensive analyses across genomic, transcriptomic, and proteomic datasets . These integrated approaches provide a more complete understanding of KIF2C's role in normal physiology and disease states.

The LinkedOmics database has been used to analyze the relationship between KIF2C expression levels and clinical-pathological characteristics of patients across multiple cancer types . This type of analysis can reveal correlations that might not be apparent when examining a single data type, such as associations between KIF2C expression and patient survival, tumor stage, or response to therapy.

Researchers can further leverage multi-omics approaches by integrating:

• Genomic data to identify mutations and copy number alterations affecting KIF2C

• Transcriptomic data to examine KIF2C expression patterns across tissues and disease states

• Proteomic data to study KIF2C protein levels and post-translational modifications

• Interactome data to identify KIF2C's binding partners in different cellular contexts

• Clinical data to correlate molecular findings with patient outcomes

Future multi-omics studies might focus on identifying biomarkers that predict response to KIF2C-targeted therapies or discovering synthetic lethal interactions that could be exploited for cancer treatment. The observation that KIF2C expression differs across cancer types suggests that tissue-specific factors influence its function, which could be elucidated through multi-omics approaches incorporating tissue-specific datasets .