Prolactin Antibody [Unconjugated]

Western blot shows lysates of mouse pituitary tissue and rat pituitary tissue. PVDF membrane was probed with 0.25 µg/mL of Goat Anti-Mouse/Rat Prolactin Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1445) followed by HRP-conjugated Anti-Goat IgG Secondary Antibody (HAF017). A specific band was detected for Prolactin at approximately 23 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

Recombinant Mouse Prolactin (1445-PL) stimulates proliferation in the Nb2-11 rat lymphoma cell line in a dose-dependent manner (orange line). Proliferation elicited by Recombinant Mouse Prolactin (10 ng/mL) is neutralized (green line) by increasing concentrations of Goat Anti-Mouse/Rat Prolactin Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1445). The ND50 is typically 0.25-1.0 µg/mL.

Prolactin was detected in immersion fixed paraffin-embedded sections of Mouse Pituitary using Goat Anti-Mouse/Rat Prolactin Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1445) at 1 µg/mL overnight at 4 °C. Tissue was stained using the Anti-Goat IgG VisUCyte™ HRP Polymer Antibody (brown; Catalog # VC004) and counterstained with hematoxylin (blue). Specific staining was localized to cytoplasm in lactotroph cells. View our protocol for IHC Staining with VisUCyte HRP Polymer Detection Reagents.

Simple Western shows lysates of mouse pituitary tissue and rat pituitary tissue, loaded at 0.2 mg/ml. A specific band was detected for Prolactin at approximately 26 kDa (as indicated) using 2.5 µg/mL of Goat Anti-Mouse/Rat Prolactin Antigen Affinity-purified Polyclonal Antibody (Catalog # af1445). This experiment was conducted under reducing conditions and using the 12-230kDa separation system.

Overexpression of ER alpha rescues miR-130a-3p-inhibited expression of PRL. The GH3 cells were transfected with the miR-130a-3p mimic and the ER alpha overexpressing vector. The rescue efficiency of ER alpha in the GH3 cells was confirmed by western blotting. (A) The protein expression level of ER alpha was detected by western blotting. GAPDH was used as loading control. (B) The protein expression level of PRL was detected by western blotting. GAPDH was used as loading control. (C) Quantitation of ER alpha protein level. Data are presented as mean ± S.E.M of n = 4 samples per group (P < 0.05 by ANOVA). (D) Quantitation of PRL protein level. Data are presented as mean ± S.E.M of n = 4 samples per group (P < 0.05 by ANOVA). Image collected and cropped by CiteAb from the following open publication (//pubmed.ncbi.nlm.nih.gov/32194503), licensed under a CC-BY license. Not internally tested by R&D Systems.

MiR-130a-3p overexpression reduces PRL expression in GH3 cells. GH3 cells were transfected with miR-130a-3p mimic and NC and then the expressions of miR-130a-3p and PRL were analyzed. (A) The expression level of miR-130a-3p was detected by quantitative real-time PCR (qRT-PCR). U6 snRNA was used to normalize the miRNA expression. Data are presented as mean ± S.E.M of n = 6 samples per group (*P < 0.05 by t-test). (B) The expression level of PRL mRNA was detected by qRT-PCR. GAPDH was used to normalize each gene expression. Data are presented as mean ± S.E.M of n = 6 samples per group (*P < 0.05 by t-test). (C) The protein level of PRL in GH3 cells was analyzed by western blotting. GAPDH was used as loading control. (D) Quantitation of the PRL protein level. Data are presented as mean ± S.E.M of n = 4 samples per group (*P < 0.05 by t-test). (E) The protein level of PRL was detected by ICC after GH3 cells transfected with miR-130a-3p mimic and NC. Scale bar, 50 μM. **P < 0.01. Image collected and cropped by CiteAb from the following open publication (//pubmed.ncbi.nlm.nih.gov/32194503), licensed under a CC-BY license. Not internally tested by R&D Systems.

Heat stress increases miR-130a-3p expression, reduces PRL and ER alpha expressions in GH3 cells. GH3 cells were separately cultured in 37 or 41°C for 24 h, after that, miR-130a-3p, PRL, and ER alpha expressions were analyzed. (A) The expression level of miR-130a-3p was detected by quantitative real-time PCR (qRT-PCR). U6 snRNA was used to normalize the miRNA expression. Data are presented as mean ± S.E.M of n = 4 samples per group (*P < 0.05 by t-test). (B) The expression levels of HSP70, ER alpha , and PRL mRNAs in GH3 cells were detected by qRT-PCR. GAPDH was used to normalize each gene expression. Data are presented as mean ± S.E.M of n = 4 samples per group (*P < 0.05, ns, not significant by t-test). (C) The protein levels of HSP70, ER alpha , and PRL in GH3 cells were analyzed by western blotting. GAPDH was used as loading control. (D) Quantitation of HSP70, ER alpha , and PRL protein levels. Data are presented as mean ± S.E.M of n = 4 samples per group (*P < 0.05 by t-test). **P < 0.01. Image collected and cropped by CiteAb from the following open publication (//pubmed.ncbi.nlm.nih.gov/32194503), licensed under a CC-BY license. Not internally tested by R&D Systems.

Heat stress reduces PRL and ER alpha expressions as well as increases miR-130a-3p expression in the pituitary gland. The mice of treated groups were placed in 40°C for 2 h each time, and the stimulus, respectively lasted 1 and 7 days. The mice in control group were fed as normal in 25°C. Then the expressions of PRL and ER alpha in the pituitary gland were analyzed. (A–B) The mRNA levels of PRL (A) and ER alpha (B) in the pituitary gland of three groups were analyzed by quantitative real-time PCR (qRT-PCR). GAPDH was used to normalize each gene expression. Data are presented as mean ± S.E.M of n = 5 animals per group. Bars that share different letter are significantly different (P < 0.05 by ANOVA). (C) The serum PRL concentration was detected by Elisa assay. Data are presented as mean ± S.E.M of n = 5 animals per group. Bars that share different letter are significantly different (P < 0.05 by ANOVA). (D) The protein levels of PRL and ER alpha in the pituitary gland were analyzed by western blotting. beta -actin was used as loading control. (E) Quantitation of PRL and ER alpha protein levels. Data are presented as mean ± SD of n = 3 animals per group. Bars that share different letter are significantly different (P < 0.05 by ANOVA). The expression levels of miR-130a-3p (F), miR-130b-3p (G), miR-301a-3p (H), and miR-301b-3p (I) were detected by qRT-PCR. U6 snRNA was used to normalize each miRNA expression. Data are presented as mean ± S.E.M of n = 5 animals per group. Bars that share different letter are significantly different (P < 0.05 by ANOVA). Image collected and cropped by CiteAb from the following open publication (//pubmed.ncbi.nlm.nih.gov/32194503), licensed under a CC-BY license. Not internally tested by R&D Systems.

• Reactivity: Mu, Rt
• Applications: WB, Simple Western, IHC, B/N, ELISA(Cap)
• Clonality: Polyclonal
• Host: Goat
• Conjugate: Unconjugated
• Concentration: LYOPH

Prolactin Antibody [Unconjugated] Summary

• Immunogen: E. coli-derived recombinant mouse Prolactin
  Leu32-Cys228
  Accession # NP_035294
• Specificity: Detects mouse and rat Prolactin in Western blots. In sandwich immunoassays, less than 1% cross-reactivity with recombinant human Prolactin is observed.
• Source: N/A
• Isotype: IgG
• Clonality: Polyclonal
• Host: Goat
• Gene: PRL
• Purity Statement: Antigen Affinity-purified
• Endotoxin Note: <0.10 EU per 1 μg of the antibody by the LAL method.

Applications/Dilutions

• Dilutions:
  • ELISA Capture (Matched Antibody Pair) 0.2-0.8 ug/mL
  • ELISA Detection (Matched Antibody Pair) 0.1-0.4 ug/mL
  • ELISA Standard (Matched Pair)
  • Immunohistochemistry 1-15 ug/mL
  • Neutralization 0.25-1.0 ug/mL
  • Simple Western 2.5 ug/mL
  • Western Blot 0.25 ug/mL
• Application Notes: ELISA Capture: Mouse Prolactin Antibody (Catalog # AF1445)
  ELISA Detection: Mouse Prolactin Biotinylated Antibody (Catalog # BAF1445)
  Standard: Recombinant Mouse Prolactin (Catalog # 1445-PL)

Packaging, Storage & Formulations

• Storage: Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
  • 12 months from date of receipt, -20 to -70 °C as supplied.
  • 1 month, 2 to 8 °C under sterile conditions after reconstitution.
  • 6 months, -20 to -70 °C under sterile conditions after reconstitution.
• Buffer: Lyophilized from a 0.2 μm filtered solution in PBS with Trehalose. See Certificate of Analysis for details.
  *Small pack size (-SP) is supplied either lyophilized or as a 0.2 µm filtered solution in PBS.
• Preservative: No Preservative
• Concentration: LYOPH
• Reconstitution Instructions: Reconstitute at 0.2 mg/mL in sterile PBS. For liquid material, refer to CoA for concentration.

Notes

This product is produced by and ships from R&D Systems, Inc., a Bio-Techne brand.

Alternate Names for Prolactin Antibody [Unconjugated]

• PRL
• Prolactin

Background

Prolactin (PRL) is a neuroendocrine pituitary hormone. Prolactin is synthesized by the anterior pituitary, placenta, brain, uterus, dermal fibroblasts, decidua, B cells, T cells, NK cells and breast cancer cells. Originally characterized as a lactogenic hormone, further studies have demonstrated broader roles in breast cancer development, regulation of reproductive function, and immunoregulation. In the immune system, Prolactin has been shown to be secreted by human PBMC and to act as a proliferative growth factor. Additionally, Prolactin treatment of human PBMC has been shown to enhance IFN-gamma production. In the breast, Prolactin-induced morphogenesis of the mammary cells is mediated through IGF-2, which in turn upregulates cyclin D1. Prolactin has several molecular forms. The predominant form is a monomer; the non-glycosylated form is 23 kDa and the glycosylated form is 25 kDa. Glycosylated Prolactin is removed from the circulation faster and has been reported to have lower biological potency. Mouse Prolactin cDNA encodes a 228 amino acid (aa) residue protein with a putative 31 aa residue signal peptide. The Prolactin receptor is a transmembrane type I glycoprotein that belongs to the cytokine hematopoietic receptor family. B cells, T cells, macrophages, NK cells, monocytes, CD34⁺ progenitor cells, neutrophils, mammary gland, liver, kidney, adrenals, ovaries, testis, prostrate, seminal vesicles, and hypothalamus have all been shown to express the Prolactin receptor. Three forms of the receptor, generated by differential splicing, have been identified. These isoforms differ in the length of their cytoplasmic domains. It is believed that the short cytoplasmic form is non-functional. Prolactin signal transduction involves the JAK/STAT families and Src kinase family (1‑9).

1. Freeman, M. et al. (2000) Physiological Reviews 80:1523.
2. Ben-Johnson, N. et al. (1996) Endoc. Rev. 17:639.
3. Cesario, T. et al. (1994) Proc. Soc. Exp. Biol. Med. 205:89.
4. Price, A.E. et al. (1995) Endoc. 136:4827.
5. Hoffmann, T. et al. (1993) J. Endoc. Invest. 16:807.
6. Cole, E. et al. (1991) Endoc. 129:2639.
7. Lewis, U. et al. (1985) Endoc. 116:359.
8. Matalk, K. (2003) Cytokine 21:187.
9. Brisken, C. et al. (2002) Dev. Cell 3:877.

Limitations

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Primary Antibodies are guaranteed for 1 year from date of receipt.

Publications for Prolactin Antibody (AF1445)(8)

Publications using AF1445

• Jinah Ha, Jungwoo Shin, Eunji Seok, Soohyun Kim, Sojung Sun, Hyunwon Yang Estradiol and progesterone regulate NUCB2/nesfatin-1 expression and function in GH3 pituitary cells and THESC endometrial cells Animal Cells and Systems 12/11/2023 [PMID: 37351263]
• Jinah Ha, Jungwoo Shin, Eunji Seok, Soohyun Kim, Sojung Sun, Hyunwon Yang Estradiol and progesterone regulate NUCB2/nesfatin-1 expression and function in GH3 pituitary cells and THESC endometrial cells Animal Cells and Systems 2023-12-11 [PMID: 37351263] (Immunocytochemistry, Rat)
• Haojie Zhang, Ting Chen, Jiali Xiong, Baoyu Hu, Junyi Luo, Qianyun Xi et al. MiR-130a-3p Inhibits PRL Expression and Is Associated With Heat Stress-Induced PRL Reduction Front Endocrinol (Lausanne) 3/3/2020 [PMID: 32194503]
• H Suzuki, T Yamamoto Chemokine CXCL14-like immunoreactivity in the &amp;alphaMSH-producing cells and PRL-producing cells of the flat-tailed house gecko pituitary J. Vet. Med. Sci., 2020-02-10;82(4):408-413. 2020-02-10 [PMID: 32037367] (IF, Gecko)
• K Handa, S Kiyohara, T Yamakawa, K Ishikawa, M Hosonuma, N Sakai, A Karakawa, M Chatani, M Tsuji, K Inagaki, Y Kiuchi, M Takami, T Negishi-Ko Bone loss caused by dopaminergic degeneration and levodopa treatment in Parkinson&#039;s disease model mice Sci Rep, 2019-09-24;9(1):13768. 2019-09-24 [PMID: 31551490] (Bioassay, Mouse)
• Christensen , Heather, Murawsky , Michael, Horseman , Nelson D, Willson , Tara A, Gregerson , Karen A Completely humanizing prolactin rescues infertility in prolactin knockout mice and leads to human prolactin expression in extrapituitary mouse tissues. Endocrinology, 2013-09-12;154(12):4777-89. 2013-09-12 [PMID: 24029242] (IHC-Fr, Mouse)
• Mak GK, Weiss S Paternal recognition of adult offspring mediated by newly generated CNS neurons. Nat. Neurosci., 2010-05-09;13(6):753-8. 2010-05-09 [PMID: 20453850] (Neutralization, Mouse)
• Korsisaari N, Ross J, Wu X, Kowanetz M, Pal N, Hall L, Eastham-Anderson J, Forrest WF, van Bruggen N, Peale FV, Ferrara N Blocking vascular endothelial growth factor-a inhibits the growth of pituitary adenomas and lowers serum prolactin level in a mouse model of multiple endocrine neoplasia type 1. Clin. Cancer Res., 2008-01-01;14(1):249-58. 2008-01-01 [PMID: 18172277] (IHC-P, Mouse)

Bioinformatics

• Gene Symbol: PRL
• Uniprot:
  • NP_035294()