The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use a concentration of 1 µg/ml.
Use a concentration of 1 µg/ml. Detects a band of approximately 76 kDa (predicted molecular weight: 74 kDa).
FunctionLamins are components of the nuclear lamina, a fibrous layer on the nucleoplasmic side of the inner nuclear membrane, which is thought to provide a framework for the nuclear envelope and may also interact with chromatin. Lamin A and C are present in equal amounts in the lamina of mammals. Play an important role in nuclear assembly, chromatin organization, nuclear membrane and telomere dynamics. Prelamin-A/C can accelerate smooth muscle cell senescence. It acts to disrupt mitosis and induce DNA damage in vascular smooth muscle cells (VSMCs), leading to mitotic failure, genomic instability, and premature senescence.
Tissue specificityIn the arteries, prelamin-A/C accumulation is not observed in young healthy vessels but is prevalent in medial vascular smooth muscle celle (VSMCs) from aged individuals and in atherosclerotic lesions, where it often colocalizes with senescent and degenerate VSMCs. Prelamin-A/C expression increases with age and disease. In normal aging, the accumulation of prelamin-A/C is caused in part by the down-regulation of ZMPSTE24/FACE1 in response to oxidative stress.
Involvement in diseaseDefects in LMNA are the cause of Emery-Dreifuss muscular dystrophy type 2 (EDMD2) [MIM:181350]. A degenerative myopathy characterized by weakness and atrophy of muscle without involvement of the nervous system, early contractures of the elbows, Achilles tendons and spine, and cardiomyopathy associated with cardiac conduction defects. Defects in LMNA are the cause of cardiomyopathy dilated type 1A (CMD1A) [MIM:115200]. Dilated cardiomyopathy is a disorder characterized by ventricular dilation and impaired systolic function, resulting in congestive heart failure and arrhythmia. Patients are at risk of premature death. Defects in LMNA are the cause of familial partial lipodystrophy type 2 (FPLD2) [MIM:151660]; also known as familial partial lipodystrophy Dunnigan type. A disorder characterized by the loss of subcutaneous adipose tissue in the lower parts of the body (limbs, buttocks, trunk). It is accompanied by an accumulation of adipose tissue in the face and neck causing a double chin, fat neck, or cushingoid appearance. Adipose tissue may also accumulate in the axillae, back, labia majora, and intraabdominal region. Affected patients are insulin-resistant and may develop glucose intolerance and diabetes mellitus after age 20 years, hypertriglyceridemia, and low levels of high density lipoprotein cholesterol. Defects in LMNA are the cause of limb-girdle muscular dystrophy type 1B (LGMD1B) [MIM:159001]. LGMD1B is an autosomal dominant degenerative myopathy with age-related atrioventricular cardiac conduction disturbances, dilated cardiomyopathy, and the absence of early contractures. LGMD1B is characterized by slowly progressive skeletal muscle weakness of the hip and shoulder girdles. Muscle biopsy shows mild dystrophic changes. Defects in LMNA are the cause of Charcot-Marie-Tooth disease type 2B1 (CMT2B1) [MIM:605588]. CMT2B1 is a form of Charcot-Marie-Tooth disease, the most common inherited disorder of the peripheral nervous system. Charcot-Marie-Tooth disease is classified in two main groups on the basis of electrophysiologic properties and histopathology: primary peripheral demyelinating neuropathy or CMT1, and primary peripheral axonal neuropathy or CMT2. Neuropathies of the CMT2 group are characterized by signs of axonal regeneration in the absence of obvious myelin alterations, normal or slightly reduced nerve conduction velocities, and progressive distal muscle weakness and atrophy. CMT2B1 inheritance is autosomal recessive. Defects in LMNA are the cause of Hutchinson-Gilford progeria syndrome (HGPS) [MIM:176670]. HGPS is a rare genetic disorder characterized by features reminiscent of marked premature aging. Note=HGPS is caused by the toxic accumulation of a mutant form of lamin-A/C. This mutant protein, called progerin, acts to deregulate mitosis and DNA damage signaling, leading to premature cell death and senescence. Progerin lacks the conserved ZMPSTE24/FACE1 cleavage site and therefore remains permanently farnesylated. Thus, although it can enter the nucleus and associate with the nuclear envelope, it cannot incorporate normally into the nuclear lamina. Defects in LMNA are the cause of cardiomyopathy dilated with hypergonadotropic hypogonadism (CMDHH) [MIM:212112]. A disorder characterized by the association of genital anomalies, hypergonadotropic hypogonadism and dilated cardiomyopathy. Patients can present other variable clinical manifestations including mental retardation, skeletal anomalies, scleroderma-like skin, graying and thinning of hair, osteoporosis. Dilated cardiomyopathy is characterized by ventricular dilation and impaired systolic function, resulting in congestive heart failure and arrhythmia. Defects in LMNA are the cause of mandibuloacral dysplasia with type A lipodystrophy (MADA) [MIM:248370]. A disorder characterized by mandibular and clavicular hypoplasia, acroosteolysis, delayed closure of the cranial suture, progeroide appearance, partial alopecia, soft tissue calcinosis, joint contractures, and partial lipodystrophy with loss of subcutaneous fat from the extremities. Adipose tissue in the face, neck and trunk is normal or increased. Defects in LMNA are a cause of lethal tight skin contracture syndrome (LTSCS) [MIM:275210]; also known as restrictive dermopathy (RD). Lethal tight skin contracture syndrome is a rare disorder mainly characterized by intrauterine growth retardation, tight and rigid skin with erosions, prominent superficial vasculature and epidermal hyperkeratosis, facial features (small mouth, small pinched nose and micrognathia), sparse/absent eyelashes and eyebrows, mineralization defects of the skull, thin dysplastic clavicles, pulmonary hypoplasia, multiple joint contractures and an early neonatal lethal course. Liveborn children usually die within the first week of life. The overall prevalence of consanguineous cases suggested an autosomal recessive inheritance. Defects in LMNA are the cause of heart-hand syndrome Slovenian type (HHS-Slovenian) [MIM:610140]. Heart-hand syndrome (HHS) is a clinically and genetically heterogeneous disorder characterized by the co-occurrence of a congenital cardiac disease and limb malformations. Defects in LMNA are the cause of muscular dystrophy congenital LMNA-related (CMD-LMNA) [MIM:613205]. It is a form of congenital muscular dystrophy. Patients present at birth, or within the first few months of life, with hypotonia, muscle weakness and often with joint contractures.
Sequence similaritiesBelongs to the intermediate filament family.
Post-translational modificationsIncreased phosphorylation of the lamins occurs before envelope disintegration and probably plays a role in regulating lamin associations. Proteolytic cleavage of the C-terminal of 18 residues of prelamin-A/C results in the production of lamin-A/C. The prelamin-A/C maturation pathway includes farnesylation of CAAX motif, ZMPSTE24/FACE1 mediated cleavage of the last three amino acids, methylation of the C-terminal cysteine and endoproteolytic removal of the last 15 C-terminal amino acids. Proteolytic cleavage requires prior farnesylation and methylation, and absence of these blocks cleavage. Sumoylation is necessary for the localization to the nuclear envelope. Farnesylation of prelamin-A/C facilitates nuclear envelope targeting.
Cellular localizationNucleus. Nucleus envelope. Farnesylation of prelamin-A/C facilitates nuclear envelope targeting and subsequent cleaveage by ZMPSTE24/FACE1 to remove the farnesyl group produces mature lamin-A/C, which can then be inserted into the nuclear lamina. EMD is required for proper localization of non-farnesylated prelamin-A/C.
Lane 1: Wild-type HAP1 cell lysate (20 µg) Lane 2: Lamin A knockout HAP1 cell lysate (20 µg) Lane 3: A431 cell lysate (20 µg) Lane 4: NIH3T3 cell lysate (20 µg) Lanes 1 - 4: Merged signal (red and green). Green - ab26300 observed at 76 kDa. Red - loading control, ab8245, observed at 37 kDa.
ab26300 was shown to recognize Lamin A when Lamin A knockout samples were used, along with additional cross-reactive bands. Wild-type and Lamin A knockout samples were subjected to SDS-PAGE. ab26300 1ug/ml and ab8245 (loading control to GAPDH) at a dilution of 1/1000 were incubated overnight at 4°C. Blots were developed with goat anti-rabbit IgG (H + L) and goat anti-mouse IgG (H + L) secondary antibodies at 1/10000 dilution for 1 hour at room temperature before imaging.
Western blot - Anti-Lamin A antibody (ab26300)This image is courtesy of an anonymous Abreview
All lanes : Anti-Lamin A antibody (ab26300) at 1/1000 dilution
Lane 1 : Mouse NIH-3T3 cells - cytosolic fraction Lane 2 : Mouse NIH-3T3 cells - nuclear fraction
Lysates/proteins at 25 µg per lane.
Secondary HRP conjugated Goat anti-rabbit at 1/5000 dilution developed using the ECL technique
All lanes : Anti-Lamin A antibody (ab26300) at 1 µg/ml
Lane 1 : Recombinant Human Lamin A protein (ab83472) at 0.1 µg Lane 2 : Recombinant Human Lamin A protein (ab83472) at 0.01 µg
Secondary Goat Anti-Rabbit IgG H&L (HRP) preadsorbed (ab97080) at 1/5000 dilution developed using the ECL technique
Performed under reducing conditions.
Predicted band size : 74 kDa
Exposure time : 10 seconds
Immunocytochemistry/ Immunofluorescence - Lamin A antibody (ab26300)This image is part of an Abreview submmited by Dr Kirk McManus.
ab26300 (1/2000) staining Lamin A in assynchronous HeLa Cells, by Immunocytochemistry/ Immunofluorescence. Secondary antibody: goat anti-rabbit conjugated to Cy3 ® (1/200). Cells counterstained with DAPI in order to highlight the nucleus.
Immunocytochemistry/ Immunofluorescence - Lamin A antibody (ab26300)This image is courtesy of an Abreview submitted by Dr Chi W Tang
ab26300 at 1/1000 staining human HeLa cells by ICC/IF. The cells were paraformaldehyde fixed, permeabilized with Triton X100 and blocked with BSA before incubation with the antibody. A Cy3 ® conjugated donkey anti-rabbit IgG was used as the secondary.
Predicted band size : 74 kDa Observed band size : 74 kDa Additional bands at : 100 kDa,45 kDa,70 kDa (possible degradation product). We are unsure as to the identity of these extra bands.
Immunocytochemistry/ Immunofluorescence - Lamin A antibody (ab26300)
ICC/IF image of ab26300 stained HeLa cells. The cells were fixed with 4% PFA (10 min) and then incubated in 1%BSA / 10% normal Goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab26300 at 1µg/ml) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 Goat anti-Rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue). This antibody also gave a positive result in 4% PFA fixed (10 min)HepG2 and MCF7 cells at 1µg/ml, and in 100% methanol fixed (5 min) HeLa, HepG2 and MCF7 cells at 1µg/ml.
References for Anti-Lamin A antibody (ab26300)
This product has been referenced in:
Mandal P et al. H3 clipping activity of glutamate dehydrogenase is regulated by stefin B and chromatin structure. FEBS J281:5292-308 (2014).
Read more (PubMed: 25263734) »
Golla U et al. Sen1p contributes to genomic integrity by regulating expression of ribonucleotide reductase 1 (RNR1) in Saccharomyces cerevisiae. PLoS One8:e64798 (2013).
Read more (PubMed: 23741394) »