Product Description
This Human Prolactin overexpression lysate was created in Baculovirus-Insect Cells and intented for use as a Western blot (WB) positive control. Purification of Prolactin protein (Cat: 10275-H08B) from the overexpression lysate was verified.
Expression Host
Baculovirus-Insect Cells
Sequence Information
A DNA sequence encoding the human PRL (CAA25214.1) (Met1-Cys227) was expressed with a polyhistidine tag at the C-terminus.
Molecule Mass
The recombinant human PRL consists of 210 amino acids and predicts a molecular mass of 24.3 kDa.
Preparation Method
Cell lysate was prepared by homogenization of the over-expressed cells in ice-cold modified RIPA Lysis Buffer with cocktail of protease inhibitors (Sigma). Cell debris was removed by centrifugation. Protein concentration was determined by Bradford assay (Bio-Rad protein assay, Microplate Standard assay). The cell lysate was boiled for 5 min in 1 x SDS loading buffer (50 mM Tris-HCl pH 6.8, 12.5% glycerol, 1% sodium dodecylsulfate, 0.01% bromophenol blue) containing 5% b-mercaptoethanol, and lyophilized.
Lysis Buffer
Modified RIPA Lysis Buffer: 50 mM Tris-HCl pH 7.4, 150 mM NaCl, 1mM EDTA, 1% Triton X-100, 0.1% SDS, 1% Sodium deoxycholate, 1mM PMSF.
Recommend Usage
1. Centrifuge the tube for a few seconds and ensure the pellet at the bottom of the tube. 2. Re-dissolve the pellet using 200μL pure water and boil for 2-5 min.
Sample Buffer
1 X Sample Buffer (1 X modified RIPA buffer+1 X SDS loading buffer).
Stability & Storage
Store at 4℃ for up to twelve months from date of receipt. After re-dissolution, aliquot and store at -80℃ for up to twelve months. Avoid repeated freeze-thaw cycles.
Application
Western Blot (WB)
Optimal dilutions/concentrations should be determined by the end user.
Prolactin (PRL) is a hormone with multiple actions in the central nervous system (CNS) spanning from physiology to pathology. PRL exerts different actions through its receptors that can be found in both neurons and glial cells (astrocytes, microglia and oligodendrocytes) of the brain.It is generally believed that in vertebrates, prolactin (PRL) is predominantly synthesized and released by pituitary lactotrophs and plays important roles in many physiological processes via activation of PRL receptor (PRLR), including water and electrolyte balance, reproduction, growth and development, metabolism, immuno-modulation, and behavior.
References
Jara LJ, et al. (2011) Prolactin and autoimmunity. Clin Rev Allergy Immunol. 40(1): 50-9. Urban A, et al. (2007) Prolactin as a factor for increased platelet aggregation. Neuro Endocrinol Lett. 28(4): 518-23. Charoenphandhu N, et al. (2007) Prolactin is an important regulator of intestinal calcium transport. Can J Physiol Pharmacol. 85(6): 569-81. Tworoger SS, et al. (2006) Prolactin and breast cancer risk. Cancer Lett. 243(2): 160-9. Ben-Jonathan N, et al. (2006) Focus on prolactin as a metabolic hormone. Trends Endocrinol Metab. 17(3): 110-6.