MC4R Antibody, Biotin conjugated
Description
Composition and Target Specificity
The MC4R Biotin-Conjugated Antibody is a rabbit-derived polyclonal antibody directed against a synthetic peptide within the N-terminal region (amino acids 10–60) of human MC4R . Key characteristics include:
Applications and Assay Performance
This antibody is validated for diverse applications, with optimized dilutions for specific assays:
In Western blotting, it detects MC4R at ~37 kDa, corresponding to its theoretical molecular weight . Competitive assays using blocking peptides confirm specificity .
Mechanistic Insights into MC4R Dysfunction
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Surface Expression Deficits: Two MC4R variants (S85G and Y268H) showed reduced membrane localization compared to wild-type receptors, as demonstrated via biotinylation assays . The antibody enabled quantification of membrane-bound MC4R, critical for assessing pathogenic obesity-linked mutations .
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Signaling Impairments: Mutant MC4R failed to activate cAMP, PKA, ERK, or CREB pathways upon α-MSH stimulation, highlighting the antibody’s utility in functional studies .
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Ligand Binding: The antibody competes with endogenous ligands like α-MSH, aiding in binding affinity studies .
Therapeutic Relevance
MC4R antibodies are pivotal in developing anti-obesity therapies. For example, biotinylated variants facilitate high-throughput drug screening by enabling streptavidin-HRP detection in ligand-receptor interaction assays .
Validation and Quality Control
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Specificity: Validated via immunoblotting with blocking peptide controls, showing complete signal abolition .
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Cross-Reactivity: Confirmed across human, mouse, and rat tissues without off-target binding .
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Lot Consistency: Rigorous batch testing ensures reproducibility in signal intensity and background levels .
Handling and Stability
Product Specs
Constituents: 50% Glycerol, 0.01M PBS, pH 7.4
Target Background
- Our research demonstrated a positive association between MC4R rs17782313 and FTO rs9939609 with zBMI, indicating a minimal contribution to childhood obesity.PMID: 29679223
- Our findings suggest that MC4R methylation status in the blood of children correlates with metabolic profiles, indicating its potential as an epigenetic biomarker for metabolic syndrome. PMID: 29598821
- This study indicates that the MC4R AA genotype is significantly associated with shorter progression-free and overall survival in glioblastoma patients undergoing radiotherapy and temozolomide treatment. PMID: 28150230
- This review focuses on the anatomical distribution of MC4R in the brain and its interactions with other pathways involved in pain modulation. PMID: 28552734
- Results indicate that the MC4R rs17782313 allele C is associated with higher HDL cholesterol and a marginal reduction in MetS risk, suggesting its potential as a biomarker for cardiometabolic risk in obese patients. PMID: 28975585
- Genotyping for the near MC4R polymorphism, rs17782313, revealed an association with moderately obese patients. PMID: 29466028
- Our study suggests that the polymorphism of FTO gene rs17817449 and GNB3 gene rs5443 (C825T) may be a genetic determinant of obesity in the Saudi population, while the impact of MC4R Asn274Ser change was not detected. PMID: 29937877
- Central melanocortin signaling exerts an influence on fat and sucrose preference in humans. PMID: 27701398
- The expression of MSH2 in patients with colon cancer may promote the expression of the obesity gene MC4R, potentially contributing to body weight gain. PMID: 28537674
- This study investigates the association of gene polymorphisms of TNF-238G/A, TNF-308G/A, IL10-1082G/A, TNFAIP3, and MC4R with comorbidity occurrence in a Romanian population with psoriasis. PMID: 29696068
- MC4R mutations are frequently observed in morbidly obese Turkish children and adolescents. PMID: 28218067
- A significant association between MC4R polymorphism and the risk of childhood obesity and BMI z-score was identified through meta-analysis. PMID: 27926527
- The observed prevalence of mutations causing impaired receptor function in this large cohort is comparable to other pediatric cohorts. MC4R deficiency tends to lead to taller stature, confirming previous clinical reports. The association of MC4R mutations with a distinct phenotype concerning metabolic characteristics remains questionable. PMID: 27654141
- The interaction between MC4R genes and dietary factors plays a significant role in the development of obesity or Type 2 diabetes phenotypes. [Review] PMID: 26666384
- This study shows that melanocortin 4 receptor constitutive activity chronically inhibits specific subtypes of neuronal voltage-gated calcium channels. PMID: 28093215
- Individuals carrying rare functional MC4R mutations exhibited a significantly higher prevalence of binge eating disorder or loss-of-control eating, independent of age, sex, and BMI. Six years after bariatric surgery, mutation carriers experienced more major complications compared to wild-type subjects, independent of age, baseline BMI, sex, operation type, and weight loss. PMID: 27222505
- FTO and near MC4R variants are associated with obesity measures in Sri Lankan populations. PMID: 26948330
- The SNP rs17782313 near MC4R gene was associated with large artery atherosclerotic stroke susceptibility in smokers. Multifactor dimensionality reduction analysis revealed a significant association between the combination of MC4R variant (rs17782313), hypertension, and smoking habit with increased risk of large artery atherosclerotic stroke in a Chinese Han population. PMID: 27701175
- In a large, all-adult, and area-based population survey, the effects of obesity-promoting minor alleles of FTO and MCR4, and their interactions with lifestyle factors, are age- and gender-related. PMID: 28384342
- The C/X genotype of the MC4R gene appears to predispose postmenopausal women to developing some metabolic syndrome symptoms. PMID: 28356726
- We conducted a candidate-gene association study in a young and athletic Italian population, testing the association of functional polymorphisms in ACE (rs4646994), FTO (rs9939609), MC4R (rs17782313), and PPARG (rs1801282) genes with body mass index (BMI) and waist-to-height ratio (WHtR). PMID: 28090739
- The family-based association analysis indicated a significant association between rs12970134 and obesity. The rs12970134 near MC4R was associated with appetite and beverage intake. PMID: 28520814
- The rs12970134 of melanocortin 4 receptor melanocortin was found to be associated with body mass index only in children with physical activity < 1h/d and sedentary behaviors >= 2h/d. PMID: 28081251
- Genetic clues shed light on the mechanism by which MC4R, FTO, and NMB influence changes in BMI and obesity. PMID: 27634552
- BDNF rs2030323 is associated with 100 to 150 greater total caloric intake per allele, with additional contributions from MC4R and, in non-Hispanic white individuals, FTO. PMID: 27551991
- The common variant rs17782313 near MC4R is associated with body mass index and measures of body fatness in children aged 2-9 years. PMID: 27583473
- An SNP in the FTO (rs9939609) gene, but not in the MC4R (rs17782313) gene, was significantly associated with prepregnancy body mass index (BMI) >/= 25 kg/m(2) (relative riskFTO = 2.1; 95% confidence interval [CI], 1.4-3.1). SNPs were not statistically associated with excessive gestational weight gain (GWG) or postpartum weight retention (PPWR). PMID: 27377581
- Genetic association studies in a population of children in Chile suggest that a polymorphism in the 3prime untranslated region of MC4R (rs17782313) is associated with eating behavior leading to overweight/obesity in the pediatric population studied (children 8-14 years old). PMID: 27730429
- FTO rs9939609, MC4R rs17782313, and PPARgamma rs1801282 polymorphisms seem to have little effect on the incidence of metabolic malfunctions and no effect on androgen-related disorders in the examined middle-aged and elderly men. PMID: 27920511
- Epigenetic alterations of the newly identified genes MC4R and HNF4a in early life might contribute to metabolic profile changes, especially increased triglyceride levels, in the cord blood of preterm infants. PMID: 27583872
- The aim of this survey is to evaluate the association of genetic variants of melanocortin-4-receptor (MC4R), pro-opiomelanocortin (POMC), apolipoprotein E (APOE), and agouti-related protein (AGRP) with obesity in the North Indian population. PMID: 26226973
- Data provide evidence that variants of MC4R and LYPLAL1 modulate body fat distribution with sexual dimorphism in a Chinese population. PMID: 26848030
- The FTO rs9939609 and MC4R rs17782313 variants may be significantly associated with nocturnal, but not daytime, blood pressure levels, and their combined effects were significant in this Chinese Han population. PMID: 26324055
- The prevalence of loss-of-function MC4R variants in obese Slovak children was 0.7%, one of the lowest frequencies in Europe. PMID: 26047380
- Carrying the A risk allele of the FTOrs9939609 and/or the C risk allele of the MC4Rrs17782313 did not influence body mass and fat mass loss, or resting energy expenditure decrease, in obese women after a 12-week energy-restricted diet intervention. PMID: 25929371
- MC4R mutations contribute to the onset of obesity. PMID: 26244670
- This molecular analysis of a family-based study showed an autosomal dominant pattern. Our study indicated that MC4R variations were found in early-onset obese Thai children, and transmission of these variations in each family followed a dominant pattern. PMID: 26782456
- The MC4R variant (rs12970134) was associated with an increased risk for obesity in Czech women. PMID: 26680479
- Uncontrolled eating scores were higher in C-allele carriers of MC4R-rs17782313 compared to non-carriers. PMID: 26588347
- Polymorphisms in the gene may be useful in identifying women at risk for osteoporosis. PMID: 25871004
- Data show that co-expression with MRAPalpha, but not MRAP2, enhances MC4R constitutive activity. MRAPalpha-enhanced MC4R constitutive activity is not dependent on MC4R complex glycosylation but may result from MRAPalpha-induced changes in MC4R conformational states. PMID: 26469516
- Drawing on recent advances in neuroscience and genetic technologies, we consider the structure and function of the melanocortin-4 receptor circuitry and its role in energy homeostasis--{REVIEW}. PMID: 26814590
- MC4R (melanocortin-4 receptor) rs17782313 gene polymorphism is associated with higher triglyceride levels in older Chinese women. PMID: 26965781
- The variant rs17782313 of MC4R is associated with an increased risk of hypertriglyceridemia in obese children and adolescents. PMID: 25948074
- Obese children carrying the FTO GG and/or the MC4R CC genotype lost more body weight in comparison to non-carriers from a lifestyle intervention. PMID: 26849546
- The interaction between FTO and MC4R polymorphisms is associated with the development of PCOS. PMID: 26032905
- The age of obesity onset in the present child generation of MC4R mutation carriers is decreasing compared to the age of onset in their parents' generation. PMID: 26238496
- FTO homozygotes and FTO heterozygotes correspond to 18.0% and 45.9% of neonates, respectively. MC4R homozygotes and MC4R heterozygotes were identified in 6.7% and 36.3% of neonates, respectively. PMID: 25503666
- There is a synergistic effect of the high-risk alleles of the FTO and MC4R genes on obesity. PMID: 25402378
- The study showed that functional variants in the MC4R gene are more commonly found in obese individuals than in non-obese individuals with binge-eating episodes. PMID: 25419636
Q&A
How should researchers validate the specificity of biotin-conjugated MC4R antibodies in different experimental models?
Validation requires a multi-step approach:
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Epitope Mapping: Confirm the antibody targets amino acids 10–60 of human MC4R using competitive ELISA with the immunizing peptide .
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Cross-Reactivity Testing: Perform Western blot (WB) on lysates from MC4R-knockout cell lines (e.g., CRISPR-edited HEK293) to rule out nonspecific bands .
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Functional Validation: Compare cAMP responses in cells transfected with wild-type vs. truncated MC4R (e.g., Q307X mutants) using FRET-based assays .
Key Validation Metrics:
What factors influence the optimal antibody concentration for MC4R detection in membrane raft vs. non-raft regions?
MC4R localizes differentially in membrane rafts, affecting antibody accessibility:
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Membrane Rafts: Use 0.55–0.75 µg/µl antibody with 1% Triton X-100 to solubilize lipid rafts .
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Non-Raft Regions: Increase concentration to 1 µg/µl and employ streptavidin-Qdot amplification for low-density targets .
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Cold Exposure: Enhance labeling efficiency by incubating cells at 4°C for 15 minutes pre-fixation to stabilize raft-localized MC4R .
How does tissue-specific glycosylation impact MC4R antibody performance in brain vs. gut studies?
MC4R undergoes tissue-specific post-translational modifications:
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Brain: Heavy sialylation reduces antibody binding affinity. Pre-treat sections with neuraminidase (1 U/ml, 37°C, 1 hr) .
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Gut: O-linked glycosylation masks the N-terminal epitope. Use enzymatic deglycosylation (PNGase F) before IHC .
What mechanisms underlie contradictory MC4R expression data in cancer models, and how can they be resolved?
Discrepancies arise from:
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Genetic Heterogeneity: BRAF-mutant tumors (e.g., HT-29) show 4× higher MC4R surface expression vs. wild-type .
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Antibody Batch Variability: Lot-to-lot differences in biotinylation efficiency (validate using HPLC-MS with a 70–80% biotinylation threshold) .
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Solution:
Can MC4R nonsense mutations (e.g., Q307X) be bypassed using pharmacological chaperones in binding studies?
Yes, but efficacy depends on mutation location:
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N-Terminal Mutants (W16X, Y35X): Treat with 100 µM gentamicin for 48 hr to induce read-through, restoring 60–70% receptor function .
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C-Terminal Mutants (Q307X): Resistant to read-through; instead, use HA-tagged MC4R constructs and anti-HA antibodies for detection .
Rescue Protocol for W16X:
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Transfect HEK293 cells with mutant MC4R.
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Treat with gentamicin (100 µM) or G418 (200 µg/ml).
How does MC4R antagonism (e.g., ML00253764) synergize with chemotherapy in BRAF-mutant cancers?
Mechanistic insights from HT-29 models :
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Apoptosis Priming: MC4R inhibition reduces Bcl-2 expression by 40%, sensitizing cells to 5-FU.
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Dosing Schedule: Administer ML00253764 (15 mg/kg bid) 24 hr before chemotherapy to maximize cAMP pathway suppression.
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Validation:
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Monitor phospho-ERK/ERK ratio (target: <0.3) via WB.
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Use Annexin V/PI co-staining to quantify early vs. late apoptosis.
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What strategies improve signal-to-noise ratios when combining MC4R antibodies with CRISPR-based MC4R reporters?
Optimize dual detection using:
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CRISPR Reporter Design: Tag endogenous MC4R with mCherry-2A peptide for red fluorescence .
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Antibody Compatibility: Use biotin-conjugated antibodies with streptavidin-655 Qdots (emission 655 nm) to avoid spectral overlap with mCherry (610 nm) .
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Sequential Staining Protocol:
Methodological Notes for Data Interpretation
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cAMP Assays: Normalize α-MSH responses to forskolin (10 µM)-induced maxima to account for G protein coupling variability .
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IHC Quantification: Use H-score (range 0–300) integrating staining intensity (0–3) and percentage of positive cells .
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Statistical Conflicts: Apply Benjamini-Hochberg correction for multi-experiment studies (FDR <5%) .
