Product datasheet
Anti-CNPase antibody [11-5B] ab6319
27 Abreviews 26 References 12 Images
Overview
Product name
Anti-CNPase antibody [11-5B]
Description
Mouse monoclonal [11-5B] to CNPase
Specificity
Reacts specifically with CNPase. In an immunoblotting assay, the antibody localizes both CNP1
(46kD) and CNP2 (48kD) bands. Immunohistochemical staining of paraffin, cryostat or
vibratome sections of rat brain reveals selective staining of oligodendrocytes in the grey and
white matter. Nerve cells and axons are not stained and astroglial cells do not appear to be
labelled.
Tested applications
IHC-FoFr, Flow Cyt, Conjugation, Dot Blot, ELISA, ICC, IHC-P, IHC-Fr, WB, ICC/IF
Species reactivity
Reacts with: Mouse, Rat, Dog, Human, Rhesus monkey
Predicted to work with: Sheep, Rabbit, Cow, Pig
Does not react with: Chicken, Guinea
pig
Immunogen
Full length native protein (purified) (Human).
Positive control
This antibody gave a positive signal in Western Blot, when tested against Human, Mouse and
Rat Spinal Cord and Brain tissue lysates. This antibody gave a positive result when used in the
following formaldehyde fixed cell lines: SKNSH IHC-P: FFPE human cerebral cortex tissue
sections.
General notes
Alternative versions available:
Anti-CNPase antibody (HRP) [11-5B] (ab201678)
Properties
Form
Liquid
Storage instructions
Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C or 80°C. Avoid freeze / thaw cycle.
Storage buffer
pH: 7.40
Preservative: 0.02% Sodium azide
Constituent: PBS
Batches contain 0.4M arginine.
Purity
IgG fraction
Clonality
Monoclonal
Clone number
11-5B
Isotype
IgG1
1
Applications
Our Abpromise guarantee covers the use of ab6319 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Application
Abreviews
Notes
IHC-FoFr
1/250. PubMed: 19259393
Flow Cyt
Use 1µg for 106 cells. ab170190-Mouse monoclonal IgG1, is suitable for use as
an isotype control with this antibody.
Conjugation
Use at an assay dependent concentration.
Dot Blot
Use at an assay dependent concentration.
ELISA
Use at an assay dependent concentration.
ICC
Use at an assay dependent concentration.
IHC-P
1/2000 - 1/1000. Perform heat mediated antigen retrieval with citrate buffer pH 6
before commencing with IHC staining protocol.
IHC-Fr
Use at an assay dependent concentration.
WB
Use a concentration of 5 µg/ml. Detects a band of approximately 48 kDa
(predicted molecular weight: 48 kDa).
ICC/IF
1/200. PubMed: 17464316
Target
Sequence similarities
Belongs to the cyclic nucleotide phosphodiesterase family.
Cellular localization
Membrane. Melanosome. Firmly bound to membrane structures of brain white matter. Identified
by mass spectrometry in melanosome fractions from stage I to stage IV.
Anti-CNPase antibody [11-5B] images
2
IHC-P image of CNPase staining on rat brain
sections using ab6319 (1:1600). heat
mediated antigen retrieval on paraffin
embedded sections was performed using
citric acid. The sections were then blocked
with 1% BSA for 10 min at 21°C. The primary
antibody was incubated for 16 hours at 21°C.
The sections were then incubated in Goat
anti-mouse (Biotin) at 1:200.
Immunohistochemistry (Formalin/PFA-fixed
paraffin-embedded sections) - Anti-CNPase
antibody [11-5B] - Oligodendrocyte Marker
(ab6319)
Carl Hobbs, King`s College London, United
Kingdom
All lanes : Anti-CNPase antibody [11-5B]
(ab6319) at 1/100 dilution
Lane 1 : Spinal Cord (Human) Tissue Lysate
- adult normal tissue (ab29188)
Lane 2 : Brain (Human) Tissue Lysate - adult
normal tissue (ab29466)
Lane 3 : Spinal Cord (Mouse) Tissue Lysate
Lane 4 : Brain (Mouse) Tissue Lysate
Lane 5 : Spinal Cord (Rat) Tissue Lysate
Lane 6 : Brain (Rat) Tissue Lysate
Western blot - Anti-CNPase antibody [11-5B] Oligodendrocyte Marker (ab6319)
Lysates/proteins at 20 µg per lane.
Secondary
Goat polyclonal to Mouse IgG - H&L - PreAdsorbed (HRP) at 1/3000 dilution
developed using the ECL technique
Performed under reducing conditions.
Predicted band size : 48 kDa
Exposure time : 1 minuteThis antibody was
raised against full length native CNPase and
is predicted to recognize both isoforms. The
predicted molecular weights of isoforms CNPI
and CNPII are 45- and 48-kDa respectively.
3
ab6319 staining CNPase in rat
oligodendrocytes by Immunocytochemistry/
Immunofluorescence.
Cells were fixed in paraformaldehyde,
blocked using 5% serum for 10 minutes at
25°C, then incubated with ab6319 at a 1/200
dilution for 2 hours at 25°C. The secondary
used was a goat anti-mouse Cy3 conjugated
polyclonal at a 1/100 dilution.
Immunocytochemistry/ Immunofluorescence CNPase antibody [11-5B] - Oligodendrocyte
Marker (ab6319)
Image courtesy of an anonymous Abreview.
Overlay histogram showing SH-SY5Y cells
stained with ab6319 (red line). The cells were
fixed with 80% methanol (5 min) and then
permeabilized with 0.1% PBS-Tween for 20
min. The cells were then incubated in 1x PBS
/ 10% normal goat serum / 0.3M glycine to
block non-specific protein-protein interactions
followed by the antibody (ab6319, 2µg/1x106
Flow Cytometry-Anti-CNPase antibody [11-5B] -
cells) for 30 min at 22ºC. The secondary
Oligodendrocyte Marker(ab6319)
antibody used was DyLight® 488 goat antimouse IgG (H+L) (ab96879) at 1/500 dilution
for 30 min at 22ºC. Isotype control antibody
(black line) was mouse IgG1 [ICIGG1]
(ab91353, 2µg/1x106 cells) used under the
same conditions. Acquisition of >5,000
events was performed. This antibody gave a
positive signal in SH-SY5Y cells fixed with 4%
paraformaldehyde (10 min)/permeabilized
with 0.1% PBS-Tween for 20 min used under
the same conditions.
4
IHC image of CNPase staining in human
cerebral cortex formalin fixed paraffin
embedded tissue section, performed on a
Leica Bond™ system using the standard
protocol F. The section was pre-treated using
heat mediated antigen retrieval with sodium
citrate buffer (pH6, epitope retrieval solution
1) for 20 mins. The section was then
incubated with ab6319, 5µg/ml, for 15 mins at
Immunohistochemistry (Formalin/PFA-fixed
room temperature and detected using an
paraffin-embedded sections) - Anti-CNPase
HRP conjugated compact polymer system.
antibody [11-5B] (ab6319)
DAB was used as the chromogen. The
section was then counterstained with
haematoxylin and mounted with DPX.
For other IHC staining systems (automated
and non-automated) customers should
optimize variable parameters such as antigen
retrieval conditions, primary antibody
concentration and antibody incubation times.
ab6319 staining CNPase in Mouse brain
tissue sections by Immunohistochemistry
(IHC-Fr - frozen sections). Tissue was fixed
with formaldehyde, permeabilized with 0.1%
Triton and blocked with mouse on mouse
blocking solution for 1 hour at 20°C. Samples
were incubated with primary antibody (1/100)
for 8 hours at 4°C. An Alexa Fluor® 488conjugated Goat anti-mouse IgG polyclonal
Immunohistochemistry (Frozen sections) - Anti-
(1/200) was used as the secondary antibody.
CNPase antibody [11-5B] (ab6319)
This image is courtesy of an Abreview submitted by
Alban Gaultier
5
ab6319 staining CNPase in the rat
oligodendrocytes by ICC/IF
(Immunoytochemistry/immunofluorescence).
Cells were fixed with paraformaldehyde,
permeabilized with methanol and blocked
with 5% BSA for 1 hour at 37°C. Samples
were incubated with primary antibody (1/100
in PBS ) for 18 hours at 4°C. An Alexa Fluor®
594-conjugated Goat anti-mouse IgG
Immunocytochemistry/ Immunofluorescence -
polyclonal (1:200) was used as the secondary
Anti-CNPase antibody [11-5B] (ab6319)
antibody.
This image is courtesy of an anonymous Abreview
All lanes : Anti-CNPase antibody [11-5B]
(ab6319) at 1/750 dilution
Lane 1 : Spinal Cord homogenate (whole
tissue lysate)
Lane 2 : Spinal Cord homogenate (whole
tissue lysate)
Lane 3 : Spinal Cord homogenate (whole
tissue lysate)
Western blot - CNPase antibody [11-5B] Oligodendrocyte Marker (ab6319)
Lysates/proteins at 2 µg per lane.
This image is courtesy of an anonymous Abreview
Secondary
HRP conjugated sheep anti-mouse IgG
Predicted band size : 48 kDa
Observed band size : 45,47 kDa
This image is courtesy of an anonymous
Abreview
6
ab6319 staining mouse brain tissue sections
by IHC-FoFr. Sections were PFA fixed and
permeabilized in 0.1% Triton X-100 prior to
blocking in 0.5% TNB for 30 minutes at 25°C.
The primary antibody was diluted 1/250 and
incubated with the sample for 18 hours at
25°C. An Alexa Fluor® 488 conjugated goat
anti-mouse antibody, diluted 1/250, was used
as the secondary.
Image demonstrates a 2-D depth projection
Immunohistochemistry (PFA perfusion fixed
through the superficial cortex.
frozen sections) - CNPase antibody [11-5B] Oligodendrocyte Marker (ab6319)
This image is courtesy of an anonymous Abreview
ab6319 staining CNPase in Dog Cerebellum
tissue sections by Immunohistochemistry
(IHC-P - paraformaldehyde-fixed, paraffinembedded sections). Tissue was fixed with
formaldehyde and blocked with 1% BSA for
10 minutes at 21°C; antigen retrieval was by
heat mediation in a citrate buffer. Samples
were incubated with primary antibody (1/1500
in blocking buffer) for 2 hours at 21°C. A
Biotin-conjugated Goat anti-mouset IgG
polyclonal (1/200) was used as the secondary
Immunohistochemistry (Formalin/PFA-fixed
antibody.
paraffin-embedded sections) - Anti-CNPase
antibody [11-5B] (ab6319)
Carl Hobbs, King`s College London, United
Kingdom
7
ICC/IF image of ab6319 stained SKNSH
cells. The cells were 4% formaldehyde fixed
(10 min) and then incubated in 1%BSA / 10%
normal goat serum / 0.3M glycine in 0.1%
PBS-Tween for 1h to permeabilise the cells
and block non-specific protein-protein
interactions. The cells were then incubated
with the antibody ab6319 at 10µg/ml
overnight at +4°C. The secondary antibody
(pseudo-colored green) was Alexa Fluor®
488 goat anti- mouse (ab150117) IgG (H+L)
Immunocytochemistry/ Immunofluorescence -
preadsorbed, used at a 1/1000 dilution for 1h.
Anti-CNPase antibody [11-5B] (ab6319)
Alexa Fluor® 594 WGA was used to label
plasma membranes (pseudo-colored red) at
a 1/200 dilution for 1h at room temperature.
DAPI was used to stain the cell nuclei
(pseudo-colored blue) at a concentration of
1.43µM for 1hour at room temperature.
ab6319 at a 1/200 dilution staining rat spinal
cord tissue sections from a 4% PFA
transcardially perfused animal by
Immunohistochemistry (Frozen sections). The
tissue was paraformaldehyde fixed and
incubated with the antibody for 18 hours.
Bound antibody was detected using an HRP
conjugated goat anti-mouse polyclonal
Immunohistochemistry (Frozen sections) CNPase antibody [11-5B] - Oligodendrocyte
Marker (ab6319)
antibody.
This image is courtesy of an Abreview
submitted by Nancy Nutile-McMenemy.
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