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

D297-3
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MONOCLONAL ANTIBODY

Anti-p75NTR (Mouse) mAb
Code No.
D297-3

Clone25-8

Subclass
Rat IgG2a

BACKGROUND: p75NTR (p75 neurotrophin receptor)
as known as LNGFR/p75NGFR/TNFRSF16/NGFR, amember of the TNF receptor superfamily, is a low affinity
nerve growth factor receptor. Expression of p75NTR isobserved in a number of non-neuronal and neuronal cells
including motor neurons during development and also indamaged neurons. It is known that p75NTR/NGF signaltransduction involved in both cell survival as well as cell
death of neural cells and apoptosis induction of neurons.

Quantity100 LConcentration
1 mg/mL

SPECIES CROSS REACTIVITY:
Species

HumanMouse

Rat

Cell

Not tested

Transfectant

Not tested

Reactivity on FCM+RELATED PRODUCT:
M081-3Rat IgG2a (isotype control) (2H3)73

PBS containing 50% glycerol, pH 7.2. No preservative iscontained.

STORAGE: This antibody solution is stable for one year
from the date of purchase when stored at -20°C.

REACTIVITY: This antibody reacts with mousePlot2: Gated by: BaF

Count145 218

FORMULATION: 100 g IgG in 100 L volume of0(clone 25-8) supernatant using protein G agarose. Thishybridoma was established by fusion of mouse myeloma
cell P3X with Wister rat lymphocyte immunized withmouse fetal hepatic cells.

290

SOURCE: This antibody was purified from hybridoma

10e0

10e1
10e2
10e3
10e4
Yellow Fluorescence (YLW-HLog)

Flow cytometric analysis of m o u s ep75NTR expressed in transfectant.
Shaded histogram indicates the reaction ofisotypic control to the cells. Open histogramindicates the reaction of D297-3 to the cells.

p75NTR on Flow cytometry.

PROTOCOL:APPLICATIONS:

Flow cytometric analysis for floating cells

Flow cytometry; 10 g/mL (final concentration)
Western blotting; Not tested
Immunoprecipitation; Not tested
Immunohistochemistry; Not tested
Immunocytochemistry; Not tested
Detailed

procedure
PROTOCOL.

isprovidedintheWe usually use Fisher tubes or equivalents as reaction tubes
for all steps described below.
1) Wash the cells 3 times with washing buffer [PBScontaining 2% fetal calf serum (FCS) and 0.09% NaN3].
*Azide may react with copper or lead in plumbing system tofollowing

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

REFERENCE:
1) Suzuki, K., et al., Gastroenterology 135, 270-281 (2008)
Clone 25-8 is used in this reference.

form explosive metal azides. Therefore, always flush plenty ofwater when disposing materials containing azide into drain.

2) Resuspend the cells with washing buffer (5x106 cells/mL).
3) Add 100 L of the cell suspension into each tube, andcentrifuge at 500 x g for 1 minute at room temperature(20~25°C). Remove supernatant by careful aspiration.
4) Add 20 L of Clear Back (human Fc receptor blocking
reagent, MBL; code no. MTG-001) to the cell pellet aftertapping. Mix well and incubate for 5 minutes at roomtemperature.
5) Add 20 L of the primary antibody at the concentration assuggested in the APPLICATIONS diluted in the washingbuffer. Mix well and incubate for 30 minutes at room
temperature.
6) Add 1 mL of the washing buffer followed bycentrifugation at 500 x g for 1 minute at room temperature.

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

D297-3
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Remove supernatant by careful aspiration.
7) Add PE conjugated anti-rat IgG antibody diluted with thewashing buffer. Mix well and incubate for 15 minutes atroom temperature.
8) Add 1 mL of the washing buffer followed bycentrifugation at 500 x g for 1 minute at room temperature.
Remove supernatant by careful aspiration.
9) Resuspend the cells with 500 L of the washing bufferand analyze by a flow cytometer.
(Positive control for Flow cytometry; Transfectant)

160530-2