Customer Review for TriMethyl-Histone H3-K56 Polyclonal Antibody (STJ29403)

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Histones are basic nuclear proteins that are responsible for the nucleosome structure of the chromosomal fiber in eukaryotes. Nucleosomes consist of approximately 146 bp of DNA wrapped around a histone octamer composed of pairs of each of the four core histones (H2A, H2B, H3, and H4). The chromatin fiber is further compacted through the interaction of a linker histone, H1, with the DNA between the nucleosomes to form higher order chromatin structures. This gene is intronless and encodes a member of the histone H3 family. Transcripts from this gene lack polyA tails; instead, they contain a palindromic termination element. This gene is located separately from the other H3 genes that are in the histone gene cluster on chromosome 6p22-p21.3. To purchase this antibody use the following link: http://www.stjohnslabs.com/trimethyl-histone-h3-k56?filter_name=STJ29403

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HOW DID OUR
ANTIBODY PERFORM?
CUSTOMER REVIEW: TriMethyl-Histone H3-K56 Polyclonal Antibody (STJ29403)

WHAT WAS USED?
Primary antibody: STJ29403. TriMethyl Histone H3-K56
Polyclonal Antibody.
Provider: St John's Laboratory
Dilution ratio: 1:250
Application: Western blot
Materials for Validation: Drosophila Schneider-2 cell line
22 kDa
16 kDa
6 kDa
Treatment of Materials: When cells were cultured to 70%-80% plating density, lysisbuffer with
protease inhibitors (Sigma) and equal volume of 2X loading buffer were added for 10 min water
bath at 85°C.

WHAT WAS THE PROTOCOL?
Gel electrophoresis: 4-15% polyacrylamide gel, constant voltage 150 V,
60 min.
Transfer: 0.2 um nitrocellulose membrane, 1 piece of gel (wet transfer),
constant voltage 100 V 60 min.
Blocking: 1X Odyssey buffer with PBS + 0.5% skimmed milk powder for
30 minutes.
Primary antibody probing: primary antibody was added to the solution of
blocking buffer, 4°C for 2 nights.
Membrane wash: 1X PBS wash for 3 times.
Secondary antibody probing: secondary antibody (Li-cor donkey anti-
rabbit) was added to the blocking buffer with 1:10000 dilution ratio, for 1 hr.
Membrane wash: 1X PBS wash for 3 times.
Visualization: Odyssey scanner.
22 kDa
16 kDa
6 kDa

WHAT WAS THE PROTOCOL?
Gel electrophoresis information：4-15% polyacrylamide gel, constant voltage 150 V, 60 min.
Transfer information：0.2 um nitrocellulose membrane, 1 piece of gel (wet transfer), constant voltage
100 V 60 min.
No. Antigen Primary antibody
Primary
antibody
dilution
ratio
Secondary
antibody
dilution
ratio
Target Band
Visualisation
Time
1
Schneider-2
nuclear lysate
STJ29403.
TriMethyl-Histone
H3-K56 Polyclonal
Antibody
1:250 1:10 000 ~17 kDa N/A
22 kDa
16 kDa
6 kDa

WHAT DID THE CUSTOMER THINK?
Testing results were provided by Serafin Colmenares from
Lawrence Berkeley National Laboratory. They stated that
“Western blot produced band of expected size in
Schneider-2 nuclear lysate.”

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Tyrosine-protein kinase that plays an essential role as cell surface receptor for neuregulins and EGF family members and regulates development of the heart, the central nervous system and the mammary gland, gene transcription, cell proliferation, differentiation, migration and apoptosis. Required for normal cardiac muscle differentiation during embryonic development, and for postnatal cardiomyocyte proliferation. Required for normal development of the embryonic central nervous system, especially for normal neural crest cell migration and normal axon guidance. Required for mammary gland differentiation, induction of milk proteins and lactation. Acts as cell-surface receptor for the neuregulins NRG1, NRG2, NRG3 and NRG4 and the EGF family members BTC, EREG and HBEGF. Ligand binding triggers receptor dimerization and autophosphorylation at specific tyrosine residues that then serve as binding sites for scaffold proteins and effectors. Ligand specificity and signaling is modulated by alternative splicing, proteolytic processing, and by the formation of heterodimers with other ERBB family members, thereby creating multiple combinations of intracellular phosphotyrosines that trigger ligand- and context-specific cellular responses. Mediates phosphorylation of SHC1 and activation of the MAP kinases MAPK1/ERK2 and MAPK3/ERK1. Isoform JM-A CYT-1 and isoform JM-B CYT-1 phosphorylate PIK3R1, leading to the activation of phosphatidylinositol 3-kinase and AKT1 and protect cells against apoptosis. Isoform JM-A CYT-1 and isoform JM-B CYT-1 mediate reorganization of the actin cytoskeleton and promote cell migration in response to NRG1. Isoform JM-A CYT-2 and isoform JM-B CYT-2 lack the phosphotyrosine that mediates interaction with PIK3R1, and hence do not phosphorylate PIK3R1, do not protect cells against apoptosis, and do not promote reorganization of the actin cytoskeleton and cell migration. Proteolytic processing of isoform JM-A CYT-1 and isoform JM-A CYT-2 gives rise to the corresponding soluble intracellular domains (4ICD) that translocate to the nucleus, promote nuclear import of STAT5A, activation of STAT5A, mammary epithelium differentiation, cell proliferation and activation of gene expression. The ERBB4 soluble intracellular domains (4ICD) colocalize with STAT5A at the CSN2 promoter to regulate transcription of milk proteins during lactation. The ERBB4 soluble intracellular domains can also translocate to mitochondria and promote apoptosis. To purchase this antibody, use the following link: http://www.stjohnslabs.com/erbb4?filter_name=STJ27886

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Ephrin receptors and their ligands, the ephrins, mediate numerous developmental processes, particularly in the nervous system. Based on their structures and sequence relationships, ephrins are divided into the ephrin-A (EFNA) class, which are anchored to the membrane by a glycosylphosphatidylinositol linkage, and the ephrin-B (EFNB) class, which are transmembrane proteins. The Eph family of receptors are divided into 2 groups based on the similarity of their extracellular domain sequences and their affinities for binding ephrin-A and ephrin-B ligands. Ephrin receptors make up the largest subgroup of the receptor tyrosine kinase (RTK) family. The protein encoded by this gene binds to ephrin-B2 and plays an essential role in vascular development. To purchase this antibody, use the following link: http://www.stjohnslabs.com/ephb4-antibody-p-65270?filter_name=STJ23550

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Core component of nucleosome. Nucleosomes wrap and compact DNA into chromatin, limiting DNA accessibility to the cellular machineries which require DNA as a template. Histones thereby play a central role in transcription regulation, DNA repair, DNA replication and chromosomal stability. DNA accessibility is regulated via a complex set of post-translational modifications of histones, also called histone code, and nucleosome remodeling. Anti-Histone H2B -http://www.stjohnslabs.com/histone-h2b- antibody-p-98699 Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation

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Plays a key role in the control of the eukaryotic cell cycle by modulating the centrosome cycle as well as mitotic onset; promotes G2-M transition, and regulates G1 progress and G1-S transition via association with multiple interphase cyclins. Required in higher cells for entry into S-phase and mitosis. Essential for early stages of embryonic development. During G2 and early mitosis, CDC25A/B/C-mediated dephosphorylation activates CDK1/cyclin complexes which phosphorylate several substrates that trigger at least centrosome separation, Golgi dynamics, nuclear envelope breakdown and chromosome condensation. Once chromosomes are condensed and aligned at the metaphase plate, CDK1 activity is switched off by WEE1- and PKMYT1-mediated phosphorylation to allow sister chromatid separation, chromosome decondensation, reformation of the nuclear envelope and cytokinesis. Anti-Cdc2 -http://www.stjohnslabs.com/cdc2-antibody-p-91597 Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation

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Peptide Dot Blotting Customer Review for Anti-Histone H2B (Tri Methyl Lys43) ...

St John's Laboratory Ltd

Histone H2B is one of the 5 main histone proteins involved in the structure of chromatin in eukaryotic cells. Histone H2B is a structural protein that helps organise eukaryotic DNA. It plays an important role in the biology of the nucleus where it is involved in the packaging and maintaining of chromosomes, regulation of transcription, and replication and repair of DNA. Histone H2B helps regulate chromatin structure and function through post-translational modifications and specialised histone variants. The antibody detects endogenous Histone H2B (Tri Methyl Lys43) protein. Anti-Histone H2B (Tri Methyl Lys43) antibody - http://www.stjohnslabs.com/histone-h2b-tri-methyl-lys43-antibody?filter_name=STJ97165 Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation

Immunofluorescence Antibody Validation Report for Anti-Actin β Antibody (STJ9...

St John's Laboratory Ltd

Actins are highly conserved proteins that are involved in various types of cell motility and are ubiquitously expressed in all eukaryotic cells. Actin participates in many important cellular processes, including muscle contraction, cell motility, cell division and cytokinesis, vesicle and organelle movement, cell signaling, and the establishment and maintenance of cell junctions and cell shape. Many of these processes are mediated by extensive and intimate interactions of actin with cellular membranes. Anti-Actin β -http://www.stjohnslabs.com/actin-b-antibody Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation

Immunohistochemistry Antibody Validation Report for Anti-Phospho-ERK 1/2 (T20...

St John's Laboratory Ltd

Serine/threonine kinase which acts as an essential component of the MAP kinase signal transduction pathway. MAPK1/ERK2 and MAPK3/ERK1 are the 2 MAPKs which play an important role in the MAPK/ERK cascade. They participate also in a signaling cascade initiated by activated KIT and KITLG/SCF. Depending on the cellular context, the MAPK/ERK cascade mediates diverse biological functions such as cell growth, adhesion, survival and differentiation through the regulation of transcription, translation, cytoskeletal rearrangements. The MAPK/ERK cascade plays also a role in initiation and regulation of meiosis, mitosis, and postmitotic functions in differentiated cells by phosphorylating a number of transcription factors. About 160 substrates have already been discovered for ERKs. Many of these substrates are localized in the nucleus, and seem to participate in the regulation of transcription upon stimulation. Anti-Phospho-ERK 1/2 (T202)-http://www.stjohnslabs.com/phospho-erk-12-t202-antibody Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation

Immunofluorescence Antibody Validation Report for Anti-β-Tubulin Antibody (ST...

St John's Laboratory Ltd

Tubulin in molecular biology can refer either to the tubulin protein superfamily of globular proteins, or one of the member proteins of that superfamily. α- and β-tubulins polymerize into microtubules, a major component of the eukaryotic cytoskeleton. Microtubules function in many essential cellular processes, including mitosis. Tubulin-binding drugs kill cancerous cells by inhibiting microtubule dynamics, which are required for DNA segregation and therefore cell division. In eukaryotes there are six members of the tubulin superfamily, although not all are present in all species (see below).[2][3] Both α and β tubulins have a mass of around 50 kDa and are thus in a similar range compared to actin with ~42 kDa. In contrast, tubulin polymers (microtubules) tend to be much bigger than actin filaments due to their cylindrical nature. Anti-β-Tubulin -http://www.stjohnslabs.com/b-tubulin-antibody-p- 98567 Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation

Immunohistochemistry Antibody Validation Report for Anti-Actin β Antibody (ST...

St John's Laboratory Ltd

Immunofluorescence Antibody Validation Report for Anti-α-tubulin Antibody (ST...

St John's Laboratory Ltd

Tubulin in molecular biology can refer either to the tubulin protein superfamily of globular proteins, or one of the member proteins of that superfamily. α- and β-tubulins polymerize into microtubules, a major component of the eukaryotic cytoskeleton. Microtubules function in many essential cellular processes, including mitosis. Tubulin-binding drugs kill cancerous cells by inhibiting microtubule dynamics, which are required for DNA segregation and therefore cell division. In eukaryotes there are six members of the tubulin superfamily, although not all are present in all species (see below).[2][3] Both α and β tubulins have a mass of around 50 kDa and are thus in a similar range compared to actin with ~42 kDa. In contrast, tubulin polymers (microtubules) tend to be much bigger than actin filaments due to their cylindrical nature. Anti-α-tubulin -http://www.stjohnslabs.com/a-tubulin-antibody-p-98572 Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation

Immunofluorescence Antibody Validation Report for Anti-AMPKα1 Antibody (STJ91...

St John's Laboratory Ltd

Catalytic subunit of AMP-activated protein kinase (AMPK), an energy sensor protein kinase that plays a key role in regulating cellular energy metabolism. In response to reduction of intracellular ATP levels, AMPK activates energy-producing pathways and inhibits energy-consuming processes: inhibits protein, carbohydrate and lipid biosynthesis, as well as cell growth and proliferation. AMPK acts via direct phosphorylation of metabolic enzymes, and by longer-term effects via phosphorylation of transcription regulators. Also acts as a regulator of cellular polarity by remodeling the actin cytoskeleton; probably by indirectly activating myosin. Anti-AMPKα1 -http://www.stjohnslabs.com/ampka1-antibody Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation

Immunofluorescence Antibody Validation Report for Anti-GAPDH Antibody (STJ96931)

St John's Laboratory Ltd

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