igfals elisa kit  (Cusabio)

Journal: eLife

Article Title: Tmem263 deletion disrupts the GH/IGF-1 axis and causes dwarfism and impairs skeletal acquisition

doi: 10.7554/elife.90949

Figure Lengend Snippet: Figure 4. TMEM263 deficiency results in marked reduction in circulating insulin-like growth factor 1 (IGF-1), IGF binding protein 3 (IGFBP3), and IGF acid labile subunit (IGFALS) levels. Serum levels of growth hormone (GH; A), IGF-1 (B), IGFBP3 (C), IGFALS (D), insulin (E), glucose (F), calcium (G), and phosphate (H) in wild-type (WT) (+/+), heterozygous (+/-), and Tmem263-KO (-/-) male and female mice at 8 weeks of age. Sample size for panel A (GH): males (WT = 9; het = 9; KO = 5) and females (WT = 10; het = 7; KO = 8). Panel B (IGF-1): males (WT = 9; het = 9; KO = 9) and females (WT = 10; het = 7; KO = 7). Panel C (IGFBP3): males (WT = 15; het = 16; KO = 11) and females (WT = 12; het = 18; KO = 10). Panel D (IGFALS): males (WT = 10; het = 10; KO = 8) and females (WT = 10; het = 10; KO = 8). Panel E (insulin): males (WT = 12; het = 17; KO = 8) and females (WT = 10; het = 16; KO = 8). Panel F (glucose): males (WT = 9; het = 14; KO = 9) and females (WT = 6; het = 15; KO = 8). Panel G and H (calcium and phosphate): males (WT = 10; het = 10; KO = 8) and females (WT = 10; het = 10; KO = 8). (I) Ratio of calcium-to-phosphate in WT, heterozygous, and KO male and female mice. Sample size for males (WT = 10; het = 10; KO = 8) and females (WT = 10; het = 10; KO = 8). All data are presented as mean ± SEM. *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001 (one-way ANOVA with Tukey’s multiple comparisons test).

Article Snippet: DOI: https://doi.org/10.7554/eLife.90949 15 of 23 Reagent type (species) or resource Designation Source or reference Identifiers Additional information Antibody Anti- phospho- STAT5 (Tyr694) (Rabbit polyclonal) Cell Signaling Technology Cat. #: sc- 137184 WB (1:500) Antibody Anti-β-Actin (Mouse monoclonal) Sigma Cat. #: A1978 WB (1:2000) Antibody Anti- TMEM263 (Rabbit polyclonal) Origene Cat. #: TA333490 WB (1:1000) Peptide, recombinant protein Recombinant human growth hormone PeproTech Cat. #: 100- 40 Dose injected into mice (3 μg/g body weight) Commercial assay or kit Pierce Cell Surface Isolation kit Thermo Fisher Scientific Cat. #: 89881 Commercial assay or kit Insulin ELISA kit Crystal Chem Cat. #: 90080 Commercial assay or kit Growth hormone (GH) ELISA kit Millipore Sigma Cat. #: EZRMGH- 45K Commercial assay or kit IGF1 ELISA kit Crystal Chem Cat. #: 80574 Commercial assay or kit IGFBP3 ELISA kit Abcam Cat. #: ab100692 Commercial assay or kit IGFALS ELISA kit Cusabio Cat. #: CSBEL011094MO Commercial assay or kit iScript cDNA synthesis kit Bio- Rad Cat. #: 1708891 Commercial assay or kit iTaq Universal SYBR Green Supermix Bio- Rad Cat. #: 1725124 Chemical compound Trizol Reagent Thermo Fisher Scientific Cat. #: 15596018 Continued

Techniques: Binding Assay

Journal: eLife

Article Title: Tmem263 deletion disrupts the GH/IGF-1 axis and causes dwarfism and impairs skeletal acquisition

doi: 10.7554/elife.90949

Figure Lengend Snippet: Figure 5. Reduced hepatic growth hormone receptor (GHR) protein level and signaling in TMEM263 knockout (KO) mice. (A) The growth hormone (GH)/insulin-like growth factor 1 (IGF-1) axis required for postnatal skeletal growth. At the onset of growth spurt, growth hormone releasing hormone (GHRH) from the hypothalamus causes the release of GH from the anterior pituitary. Circulating GH binds to its receptor (GHR) in liver and other peripheral tissues to induce the synthesis and secretion of IGF-1, which then acts in an endocrine, paracrine, and/or autocrine manner to induce skeletal growth. (B) Expression levels of Ghr (growth hormone receptor), Igf1, Igfals (IGF binding protein acid labile subunit), and Igfbp3 (IGF binding protein 3) transcripts in the liver of wild-type (WT) and KO mice. Sample size of male mice (WT, n=8; KO, n=8) and female mice (WT, n=8; KO, n=8). (C) Immunoblot analysis of GHR protein levels in the liver of WT (n=7) and KO (n=7) mice. Molecular weight markers are indicated on the left. (D) Quantification of the immunoblot results as shown in C (n=7 per genotype). (E–F) Reduced hepatic GH-induced signaling in KO (-/-; n=5) mice relative to WT (+/+; n=4) controls. Immunoblot analysis of phospho-JAK2 (Tyr1008), total JAK2, phospho-STAT5 (Y694), and total STAT5 in liver lysates from control male mice not injected with GH (E) and male mice injected with recombinant GH (F). Molecular weight markers are indicated on the left of the gel. (G) Quantification of the immunoblot results as shown in F (WT, n=4; KO, n=5). All data are presented as mean ± SEM. **p<0.01; ***p<0.001; ****p<0.0001 (one-way ANOVA with Tukey’s multiple comparisons test for data in B and two-tailed Student’s t-test for data in D and F).

Article Snippet: DOI: https://doi.org/10.7554/eLife.90949 15 of 23 Reagent type (species) or resource Designation Source or reference Identifiers Additional information Antibody Anti- phospho- STAT5 (Tyr694) (Rabbit polyclonal) Cell Signaling Technology Cat. #: sc- 137184 WB (1:500) Antibody Anti-β-Actin (Mouse monoclonal) Sigma Cat. #: A1978 WB (1:2000) Antibody Anti- TMEM263 (Rabbit polyclonal) Origene Cat. #: TA333490 WB (1:1000) Peptide, recombinant protein Recombinant human growth hormone PeproTech Cat. #: 100- 40 Dose injected into mice (3 μg/g body weight) Commercial assay or kit Pierce Cell Surface Isolation kit Thermo Fisher Scientific Cat. #: 89881 Commercial assay or kit Insulin ELISA kit Crystal Chem Cat. #: 90080 Commercial assay or kit Growth hormone (GH) ELISA kit Millipore Sigma Cat. #: EZRMGH- 45K Commercial assay or kit IGF1 ELISA kit Crystal Chem Cat. #: 80574 Commercial assay or kit IGFBP3 ELISA kit Abcam Cat. #: ab100692 Commercial assay or kit IGFALS ELISA kit Cusabio Cat. #: CSBEL011094MO Commercial assay or kit iScript cDNA synthesis kit Bio- Rad Cat. #: 1708891 Commercial assay or kit iTaq Universal SYBR Green Supermix Bio- Rad Cat. #: 1725124 Chemical compound Trizol Reagent Thermo Fisher Scientific Cat. #: 15596018 Continued

Techniques: Knock-Out, Expressing, Binding Assay, Western Blot, Molecular Weight, Control, Injection, Recombinant, Two Tailed Test