IGF1 N15 Human

What is IGF1 N15 Human and how is it produced?

IGF1 N15 Human Recombinant is a single, non-glycosylated polypeptide chain containing 70 amino acids with a molecular mass of 7.74kDa . It is produced in Escherichia coli expression systems and purified using proprietary chromatographic techniques . The N15 designation indicates nitrogen-15 stable isotope labeling throughout the protein structure, which creates a mass shift while maintaining identical chemical and biological properties to unlabeled IGF1.

The amino acid sequence is: GPETLCGAEL VDALQFVCGD RGFYFNKPTG YGSSSRRAPQ TGIVDECCFR SCDLRRLEMY CAPLKPAKSA .

This labeled variant is particularly valuable for mass spectrometry applications, metabolic tracing studies, and experiments requiring distinction between endogenous and exogenous IGF1.

What are the physiological roles of IGF1 in human development and metabolism?

IGF1 (Insulin-like growth factor-1) functions as the main mediator of human growth hormone (HGH) and plays multifaceted roles in human physiology:

• Growth and development: IGF1 stimulates bone and tissue growth, promotes lean muscle mass development, and is essential for normal skeletal myogenesis .

• Cellular function: IGF1 induces various cellular responses including proliferation, differentiation, migration, and survival .

• Muscle development: IGF1 increases satellite cell proliferation and myoblast proliferation/differentiation during normal growth or regeneration after skeletal muscle injury .

• Metabolic regulation: It stimulates glucose transport at much lower concentrations than insulin and plays a role in glucose and lipid metabolism .

• Neuroprotection: IGF1 and its metabolites demonstrate protective effects in hypoxic-ischemic conditions .

IGF1 is one of the most potent activators of the AKT signaling pathway, which stimulates proliferation and inhibits programmed cell death .

How do IGF1 levels change throughout human development?

IGF1 levels follow a characteristic pattern across the human lifespan:

• Infancy: Levels are relatively low

• Childhood: Gradual increase occurs during childhood

• Puberty: Levels peak during puberty when growth hormone secretion is highest

• Adulthood: Progressive decline with advancing age

• Pregnancy: Serum IGF1 increases approximately 2-fold

These normal fluctuations must be considered when interpreting IGF1 measurements in research and clinical settings. Abnormal IGF1 levels may indicate conditions such as growth hormone deficiency or excess, nutritional deficiencies, or metabolic disorders .

How should IGF1 N15 Human be handled and reconstituted for optimal experimental results?

Storage recommendations:

• Store lyophilized IGF1 N15 desiccated below -18°C for long-term stability (though it remains stable at room temperature for up to 3 weeks)

• After reconstitution, store at 4°C if using within 2-7 days

• For longer storage post-reconstitution, maintain below -18°C

• Avoid repeated freeze-thaw cycles to prevent protein degradation

Reconstitution protocol:

• Reconstitute lyophilized IGF1 N15 in sterile 18MΩ-cm H₂O at a concentration not less than 100μg/ml

• For long-term storage of reconstituted protein, add a carrier protein (0.1% HSA or BSA)

• Solution can be further diluted in appropriate buffers depending on experimental requirements

Quality control considerations:

• Verify purity via RP-HPLC and SDS-PAGE (should exceed 97.0%)

• Confirm biological activity through cell proliferation assays using serum-free human MCF-7 cells (ED50 should be less than 2ng/ml, corresponding to >5.0×10⁵ IU/mg)

How does cyclic glycine-proline (cGP) regulate IGF1 function in experimental models?

Cyclic glycine-proline (cGP) represents a novel auto-regulatory mechanism for IGF1 function that has significant research implications:

Regulatory mechanism:

• cGP is a metabolite of IGF1 that demonstrates bidirectional regulation of IGF1 activity

• It promotes IGF1 activity when IGF1 is insufficient but inhibits IGF1 activity when IGF1 is excessive

• Mathematical modeling reveals that cGP modulates IGF1 effect by altering the binding of IGF1 to its binding proteins

• This mechanism dynamically regulates the balance between bioavailable and non-bioavailable IGF1

Experimental evidence:

• In rats, cGP improves recovery from ischemic brain injury

• In mice, cGP inhibits the growth of lymphomic tumors

• Cell culture experiments support the bidirectional regulatory effect

This represents a significant advancement in understanding IGF1 homeostasis and has potential therapeutic implications for conditions characterized by either insufficient or excessive IGF1 activity.

What methodologies can distinguish between IGF1 and insulin signaling in research models?

Despite structural and functional similarities between IGF1 and insulin, several methodological approaches can differentiate their signaling:

Concentration-response differentiation:

• IGF1 typically shows effects at much lower concentrations compared to insulin for certain responses (e.g., glycogen synthesis, DNA synthesis, glucose uptake)

• Carefully designed dose-response experiments can identify the primary pathway involved

Receptor-specific approaches:

• Use of IGF1R-specific or insulin receptor-specific blocking antibodies

• Selective small molecule inhibitors of each receptor type

• Genetic models with receptor knockdown/knockout

Downstream signaling analysis:

• While both activate PI3K/AKT and MAPK pathways, the relative activation levels and kinetics differ

• Phosphoproteomic analysis can identify differential pathway activation patterns

• Time-course experiments can reveal signaling dynamics unique to each hormone

Experimental design considerations:

• Cross-reactivity between receptors occurs at higher concentrations

• Hybrid insulin/IGF1 receptors may complicate interpretation

• Receptor expression levels vary between cell types and influence relative responsiveness

What are the best experimental models for studying IGF1-induced muscle hypertrophy?

Research on IGF1's effects on muscle development employs several validated models:

Cellular models:

• Human primary myoblast cultures allow examination of both proliferation and differentiation processes

• Serum-free culture systems help isolate IGF1-specific effects

Key measurements in myotube studies:

• Mean number of nuclei per myotube

• Fusion index (percentage of nuclei in myotubes)

• Myosin heavy chain (MyHC) content

• Myotube diameter and cross-sectional area

Mechanisms of IGF1-induced hypertrophy:
Research has revealed multiple pathways:

• Enhancement of myoblast proliferative lifespan (though this effect is more limited in human compared to rodent cells)

• Delayed replicative senescence in human myoblasts

• Non-proliferative recruitment of mononucleated cells

• Recruitment of reserve cells in human skeletal muscle

The most comprehensive studies combine multiple approaches to distinguish between these mechanisms.

How can IGF1 N15 Human be utilized in proteomic and metabolic studies?

The stable isotope labeling in IGF1 N15 makes it particularly valuable for various analytical applications:

Mass spectrometry applications:

• Serves as an internal standard for absolute quantification (AQUA) of endogenous IGF1

• The mass shift created by N15 incorporation enables differentiation between endogenous and exogenous IGF1 in complex samples

• Facilitates studies of IGF1 post-translational modifications

Metabolic tracing studies:

• Enables tracking of nitrogen transfer from IGF1 to downstream metabolites

• Allows investigation of IGF1 degradation pathways

• Facilitates studies of amino acid recycling from IGF1

Experimental design considerations:

• Background N15 levels should be accounted for in experimental design

• Sample preparation methods must preserve the integrity of the labeled protein

• Appropriate controls should include unlabeled IGF1 to account for non-isotope-related effects

What methods are most effective for studying IGF1 homeostasis in relation to disease models?

IGF1 homeostasis has significant implications for various disease states:

Measurement approaches:

• Total IGF1 quantification via validated immunoassays

• Free (unbound) IGF1 measurement to assess bioavailable fraction

• IGF binding protein (IGFBP) quantification to understand regulatory mechanisms

• IGF1 receptor activation assessment via phosphorylation status

Disease-specific methodological considerations:

For neurodegenerative disorders:

• Low IGF1 levels have been linked to Alzheimer's disease

• Requires assessment of both peripheral and central nervous system IGF1 levels

• Often necessitates correlation between circulating IGF1 and neurological outcomes

For muscle-wasting conditions:

• IGF1 plays an important role in preventing muscle atrophy

• Research typically includes measurement of muscle-specific IGF1 isoforms

• Analysis of satellite cell activation and protein synthesis/degradation balance

For growth disorders:

• IGF1 testing helps evaluate GH deficiency or excess

• IGF1 provides a stable indicator of average GH levels, avoiding the pulsatile nature of GH secretion

• Growth studies should include age-matched controls due to the significant variation in normal IGF1 levels across development

How should researchers design experiments to account for IGF1 binding proteins?

IGF1 homeostasis is dynamically regulated through reversible binding to circulating and tissue-associated IGF binding proteins (IGFBPs) , which significantly impacts experimental design:

Experimental approaches to account for IGFBPs:

• Measure both total and free IGF1 in biological samples

• Consider using des-(1-3) IGF-1 (des-IGF-1), which has reduced affinity for IGFBPs

• Include IGFBP measurements alongside IGF1 quantification

• Use defined media without IGFBPs for cell culture experiments

Impact of cGP on IGF1-IGFBP interactions:

• cGP can normalize IGF1 function by changing the binding of IGF1 to its binding proteins

• This mechanism dynamically regulates the balance between bioavailable and non-bioavailable IGF1

• Experiments investigating IGF1 function should consider the potential presence and impact of endogenous cGP

What quality control parameters should be evaluated when working with IGF1 N15 Human?

Ensuring consistency and reliability in IGF1 N15 Human experiments requires rigorous quality control:

Physical and chemical parameters:

• Purity assessment via RP-HPLC and SDS-PAGE (>97% purity required)

• Protein concentration verification

• Mass spectrometry confirmation of N15 incorporation

• Structural integrity verification

Biological activity verification:

• Cell proliferation assays using serum-free human MCF-7 cells (ED50 <2ng/ml)

• Specific activity calculation (should exceed 5.0×10⁵ IU/mg)

• Receptor activation assessment via phosphorylation assays

• Comparison to unlabeled IGF1 standards to ensure equivalent potency

Storage and stability monitoring:

• Activity assessment after storage periods

• Evaluation after reconstitution

• Monitoring of activity retention through freeze-thaw cycles

• Temperature stress testing

How can researchers accurately interpret IGF1 measurements in complex biological systems?

Interpreting IGF1 data requires consideration of numerous biological variables:

Age-dependent reference ranges:

• IGF1 levels vary significantly with age, peaking during puberty and declining in adulthood

• Age-matched controls are essential for valid comparisons

• Developmental stage must be considered when interpreting results

Physiological confounders:

• Nutritional status significantly affects IGF1 levels

• Growth hormone pulsatility influences IGF1 production

• Pregnancy increases IGF1 levels approximately 2-fold

• Chronic kidney or liver diseases can alter IGF1 metabolism

Analytical considerations:

• Distinction between total and free (bioavailable) IGF1

• IGFBP levels affect the proportion of bioavailable IGF1

• Cross-reactivity with IGF2 in some assay systems

• Potential interference from heterophilic antibodies