HA722216

Phospho-MCM2 (S40) Recombinant Rabbit Monoclonal Antibody [JE35-01]

cat.: HA722216

Product Type:	Recombinant Rabbit monoclonal IgG, primary antibodies
Species reactivity:	Human, Mouse, Rat
Applications:	WB, IF-Cell, IHC-P, FC, IF-Tissue
Clonality:	Monoclonal
Clone number:	JE35-01

Form:	Liquid
Storage condition:	Store at +4℃ after thawing. Aliquot store at -20℃. Avoid repeated freeze / thaw cycles.
Storage buffer:	1*TBS (pH7.4), 0.05% BSA, 40% Glycerol. Preservative: 0.05% Sodium Azide.
Concentration:	1ug/ul
Purification:	Protein A affinity purified.
Molecular weight:	Predicted band size: 102 kDa
Isotype:	IgG
Immunogen:	Synthetic phosphopeptide corresponding to residues surrounding Ser40 of human MCM2.
Positive control:	HeLa cell lysate, 293T cell lysate, NIH/3T3 cell lysate, C6 cell lysate, HeLa, human tonsil tissue, mouse spleen tissue, rat spleen tissue.
Subcellular location:	Nucleus, Chromosome.
Recommended Dilutions: WB IF-Cell IHC-P FC IF-Tissue	1:2,000 1:1,000 1:1,000 1:1,000 1:200
Uniprot #:	SwissProt: P49736 Human \| P97310 Mouse Entrez Gene: 312538 Rat
Alternative names:	BM28 CCNL 1 CCNL1 CDC like 1 cdc19 CDCL 1 CDCL1 Cell devision cycle like 1 Cyclin like 1 D3S3194 DNA replication licensing factor MCM2 KIAA0030 MCM 2 MCM2 MCM2 minichromosome maintenance deficient 2 mitotin MCM2 minichromosome maintenance deficient 2 mitotin (S. cerevisiae) MCM2 minichromosome maintenance deficient 2, mitotin MCM2_HUMAN MGC10606 Minichromosome maintenance complex component 2 Minichromosome maintenance deficient 2 (mitotin) Minichromosome maintenance deficient 2 mitotin Minichromosome maintenance protein 2 Minichromosome maintenance protein 2 homolog Mitotin Nuclear protein BM28 OTTHUMP00000216047 OTTHUMP00000216050

Images

Fig1: Western blot analysis of Phospho-MCM2 (S40) on different lysates with Rabbit anti-Phospho-MCM2 (S40) antibody (HA722216) at 1/2,000 dilution.

Lane 1: HeLa cell lysate
Lane 2: 293T cell lysate
Lane 3: NIH/3T3 cell lysate
Lane 4: C6 cell lysate
Lane 5: 293T cell lysate, the membrane treated with λpp for 1 hour

Lysates/proteins at 20 µg/Lane.

Predicted band size: 102 kDa
Observed band size: 130 kDa

Exposure time: 14 seconds; ECL: K1801;

4-20% SDS-PAGE gel.

Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (HA722216) at 1/2,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.

Fig2: Immunocytochemistry analysis of HeLa cells treated with or without λpp labeling Phospho-MCM2 (S40) with Rabbit anti-Phospho-MCM2 (S40) antibody (HA722216) at 1/1,000 dilution.

Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-Phospho-MCM2 (S40) antibody (HA722216) at 1/1,000 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.

Beta tubulin (M1305-2, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution.

	Fig3: Immunohistochemical analysis of paraffin-embedded human tonsil tissue with Rabbit anti-Phospho-MCM2 (S40) antibody (HA722216) at 1/1,000 dilution. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH₂O and PBS, and then probed with the primary antibody (HA722216) at 1/1,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
	Fig4: Immunohistochemical analysis of paraffin-embedded mouse spleen tissue with Rabbit anti-Phospho-MCM2 (S40) antibody (HA722216) at 1/1,000 dilution. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH₂O and PBS, and then probed with the primary antibody (HA722216) at 1/1,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
	Fig5: Immunohistochemical analysis of paraffin-embedded rat spleen tissue with Rabbit anti-Phospho-MCM2 (S40) antibody (HA722216) at 1/1,000 dilution. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH₂O and PBS, and then probed with the primary antibody (HA722216) at 1/1,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

Fig6: Flow cytometric analysis of HeLa cells labeling Phospho-MCM2 (S40).

Cells were fixed and permeabilized. Then stained with the primary antibody (HA722216, 1μg/mL) (red) compared with Rabbit IgG Isotype Control (green). After incubation of the primary antibody at +4℃ for an hour, the cells were stained with a iFluor™ 488 conjugate-Goat anti-Rabbit IgG Secondary antibody (HA1121) at 1/1,000 dilution for 30 minutes at +4℃. Unlabelled sample was used as a control (cells without incubation with primary antibody; black).

Note: All products are “FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE”.