Anti-Calmegin antibody, rabbit serum
Product code73-034

Size

100 µlStorage

Store 4℃ for short term For long term storage store at -20℃.
Aliquot to avoid repeated freezing and thawing.

Concentration

N/A

Buffer

0.1% sodium azide

PurityRabbit antiserum

ImmunogenC-terminal peptide of mouse Calmegin, DESPGSGDAPLKSLRKRRVRKD,
conjugated with KLH

IsotypeRabbit IgG

Reactivity

Mouse

Special notes

Validation: Specificity validated with knock-out mouse (Fig.2.)

Application

1. Western blotting (1/1,000 dilution)
2. Immunoprecipitation (1/100~1/1,000 dilution).
3. Immunofluorescent staining (1/100~1/1,000 dilution)
4. Immunohistochemistry (Paraffin embedded) (1/1,000 dilution)

Background

Calmegin plays an important role in sperm fertility. Binds calcium ions.
Specifically expressed during male meiotic germ cell development.
Molecular mass: 69,431 with 611 amino acids.

Data Link

uniprot/P52194 mouse Calmegin ,Gene ID 12745 mouse Clgn

Please note: All products are FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC
PROCEDURES. NOT FOR MILITARY USE.
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Data Images: 73-034 Anti-Calmegin antibody, rabbit serum

Fig.1 Western blot analysis of Calmegin

in lysates of mouse testis and sperm.

Proteins in the lysates (10 µg) were separated on SDS-PAGE (10~20% gel) and blotted to PVDF
membrane. It was reacted with anti-Calmegin-antibody at 1/1,000 dilution.
As the second antibody, goat anti-rabbit IgG conjugated with HRP (Abcam; ab97051) was used at1/10,000 dilution.

Fig.2 Western blotting analysis of testis extracts of wild-type and knockout mice with anti-Calmeginantibody.
20 μg of Triton X-100 extracts from mouse testes was reacted with anti-Calmedin antiserum at 1/1,000
dilution. Arrow indicates the position of intact Calmegin.
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Fig.3. Immunoprecipitation of Calmegin from mouse testis.
One mg of testis lysate was incubated with 2 μl of anti-Calmegin antiserum and 50 μl. of protein-A
conjugated magnetic beads (Miltenyi Biotec) and immunoprecipitated according to the protocol ofsupplier. The immunoprecipitated sample was analyzed by western blotting with the antibody at
1/1,000 dilution.
1. Input

2. non-immune serum 3. Anti-Calmegin antiserum

Fig.4 Immunofluorescent staining of Calmegin in NIH3T3 cells with anti-Calmegin antibody
Fixation of the cells in 4% paraaformaldehyde over night
Permeabilization in 0.25% Triton X-100/PBS for 10 min
Blocking in 1.5% BSA/PBS for 30 min
1st antibodies 1/100 diluted by blocking buffer over night
2nd Goat anti rabbit IgG conjugated with Alex 488 (1:1000 dilution) for 60min
Nuclei were stained with DAPI

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Fig.5 Immunohistological staining of Calmegin in testis section with anti-Calmegin antibody
Deparaffinization

LemosolRA (#122-03991,Wako, Osaka)

Rehydration
Antigen retrieval

Histo/Zyme (Cat.# k046; Diagnostic BioSystems)

Washing

PBST (0.25% triton X-100/PBS-)

Blocking

10 % FBS / PBST

1st antibody1/1,000 dilution in PBS- 4℃ O/NWashingPBS-

2nd antibody

Goat anti rabbit IgG conjugated with Alex 488 (1:1000 dilution) for 60min

WashingPBS-

5 min, 3 times

DAPI

1.0μg/mL DAPI in TBS

Washing

PBS-

Mount

ImmunoSelect Antifading Mounting Medium (SCR-38447; Dianova

30 min10 min4 / 4

BioAcademia,Inc. Tel. 81-6-6877-2335

Fax. 81-6-6877-2336 info@bioacademia.co.jp https://www.bioacademia.co.jp/en/