M084-4
Page 1 of 2

For Research Use Only.
Not for use in diagnostic procedures.

MONOCLONAL ANTIBODY

Anti-Podocalyxin (PCLP1) (Human) mAb-FITC
Code No.
M084-4

Clone
53D11Subclass
Mouse IgG2a

BACKGROUND: Recent studies with avian embryos
and murine embryonic stem cells have suggested that
hematopoietic cells are derived from hemangioblasts, thecommon precursors of hematopoietic and endothelial cells.
Hara et al. molecularly cloned podocalyxin-like protein 1(PCLP1) as a novel surface marker for endothelial-likecells in the AGM (aorta-gonad-mesonephros) region ofmouse embryos, where long-term repopulatinghematopoietic stem cells (LTR-HSCs) are known to arise.
PCLP1+ CD45- cells in the AGM region incorporated
acetylated low-density lipoprotein and produced both
hematopoietic and endothelial cells when cocultured with
OP9 stromal cells. Moreover, multiple lineages ofhematopoietic cells were generated in vivo when PCLP1+
CD45- cells were injected into neonatal liver ofbusulfan-treated mice. Today it is reported that the PCLP1
is identical with the Podocalyxin.

SOURCE: This antibody was purified from mouse
ascites fluid using protein A agarose beads. Thishybridoma (53D11) was established by fusion of mouse
myeloma cell P3U1 with Balb/c mouse splenocyte
immunized with CHO cell expressing full-length human
Podocalyxin (PCLP1).

FORMULATION: 50 µg IgG in 100 µL volume ofPBS containing 1% BSA and 0.09% NaN 3 .

Quantity
100 µL

Concentration
500 µg/mL

SPECIES CROSS REACTIVITY:
Species

Human

Mouse

RatCells

HUVEC, MCF7

Not tested

Not tested

Reactivity on FCM+REFERENCES:1) Kuhara, M., et al., Cell Struct. Funct. 34, 23-30 (2009) [FCM]
2) Doyonnas, R., et al., Blood 105, 4170-4178 (2005)
3) Minegishi, N., et al., Blood 102, 896-905 (2003)
4) Schopperle, M.W., et al., Biochem. Biophys. Res. Commun.
300, 285-290 (2003)
5) Minehata, K., et al., Blood 99, 2360-2368 (2002)
6) Doyonnas, R., et al., J. Exp. Med. 194, 13-27 (2001)
7) Hara, T., et al., Immunity 11, 567-578 (1999)
8) Kershaw, B.D., et al., J. Biol. Chem. 272, 15708-15714 (1997)
This antibody is used in reference number 1).

RELATED PRODUCTS:
D072-3 Anti-Podocalyxin (PCLP1) (Mouse) mAb (10B9)
D072-4 Anti-Podocalyxin (PCLP1) (Mouse) mAb-FITC (10B9)
D072-5 Anti-Podocalyxin (PCLP1) (Mouse) mAb-PE (10B9)
D072-6 Anti-Podocalyxin (PCLP1) (Mouse) mAb-Biotin (10B9)
M084-3 Anti-Podocalyxin (PCLP1) (Human) mAb (53D11)
M084-5 Anti-Podocalyxin (PCLP1) (Human) mAb-PE (53D11)
M085-3 Anti-Podocalyxin (PCLP1) (Human) mAb (4H11)*Azide may react with copper or lead in plumbing system toform explosive metal azides. Therefore, always flush plenty ofwater when disposing materials containing azide into drain.

STORAGE: This antibody solution is stable for one year
from the date of purchase when stored at 4°C.

REACTIVITY: This antibody reacts with human
Podocalyxin (PCLP1) on Flow cytometry.

APPLICATION:

Flow cytometry; 30 µg/mL (final concentration)*Please refer to the data sheet (MBL code no. M084-3)
for other applications.
Detailed procedure is provided in thefollowing

PROTOCOL.

Flow cytometric analysis of Podocalyxin/
PCLP1 expression on HUVEC cells (left)
and MCF7 cells (right). Open histograms
indicate the reaction of isotypic control tothe cells. Shaded histograms indicate thereaction of M084-4 to the cells.
PROTOCOL:
Flow cytometric analysis for adherent cells

INTENDED USE:
For Research Use Only. Not for use in diagnostic procedures.

We usually use Fisher tubes or equivalents as reaction tubes
for all steps after 2).
1) Detach the cells from culture dish by using cell

MEDICAL & BIOLOGICAL LABORATORIES CO., LTD.
URL http://ruo.mbl.co.jp
e-mail support@mbl.co.jp, TEL 052-238-1904

M084-4
Page 2 of 2

dissociation buffer (Invitrogen; code no. 13151-014).
2) Wash the cells 3 times with washing buffer [PBScontaining 2% fetal calf serum (FCS) and 0.09% NaN 3 ].
3) Resuspend the cells with washing buffer (5x106cells/mL).
4) Add 50 µL of the cell suspension into each tube, andcentrifuge at 500 x g for 1 minute at room temperature.
Remove supernatant by careful aspiration.
5) Add 10 µL of normal goat serum containing 1 mg/mL
normal human IgG and 0.09% NaN 3 to the cell pellet
after tapping. Mix well and incubate for 5 minutes atroom temperature.
6) Add 20 µL of the primary antibody as suggested in theAPPLICATIONS. Mix well and incubate for 30
minutes at room temperature.
7) Add 1 mL of the washing buffer followed bycentrifugation at 500 x g for 1 minute at roomtemperature. Remove supernatant by careful aspiration.
8) Resuspend the cells with 500 µL of the washing bufferand analyze by a flow cytometer.
(Positive controls for Flow cytometry; HUVEC and
MCF7)160620-5