Product datasheet
Anti-beta 1 Spectrin antibody [4C3] ab2808
6 References 11 Images
Overview
Product name
Anti-beta 1 Spectrin antibody [4C3]
Description
Mouse monoclonal [4C3] to beta 1 Spectrin
Specificity
Detects spectrin from erythrocytes, brain and muscle cells. This antibody has been shown to
specifically detect the two known alternatively spliced forms of spectrin, beta-1 epsilon-1,
present in erythrocytes, and beta-1 epsilon-2, present in nerve and striated muscle cells. It does
not cross-react with alpha-2 spectrin or either of the fodrin subunits.
Tested applications
Flow Cyt, IHC-P, ICC/IF, WB, IHC-P
Species reactivity
Reacts with: Mouse, Rat, Human
Immunogen
Full length native protein (purified) corresponding to Human beta 1 Spectrin. Purified human
erythrocyte beta-1 spectrin.
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
Preservative: 0.05% Sodium azide
Constituent: PBS
Purity
Ascites
Primary antibody notes
Spectrin (Sp), the most abundant of the erythrocyte membrane skeleton proteins, helps these
cells maintain their characteristic biconcave shape while remaining flexible and elastic.
Erythrocyte Sp is a heterodimer composed of a 280 kDa alpha subunit and a 246 kDa beta
subunit which associate in a side-to-side, antiparallel configuration to form a 100 nm rod-like
structure. Sp in other tissues may be composed of distinct but homologous alpha and beta
subunits, sometimes referred to as fodrin. A newly introduced nomenclature designates the Sp
subunits of the erythrocyte as alpha-1 and beta-1, and the fodrin subunits as alpha-2 and beta-2.
Alternatively spliced forms of each are designated as epsilon-1, epsilon-2, etc. (e.g. beta-1
epsilon-1).
Clonality
Monoclonal
Clone number
4C3
Isotype
IgG1
Applications
1
Our Abpromise guarantee covers the use of ab2808 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
Flow Cyt
1/50.
IHC-P
1/200. Perform heat mediated antigen retrieval before commencing with IHC
staining protocol.
ICC/IF
1/10 - 1/200.
WB
1/100.
IHC-P
1/20 - 1/200.
Perform antigen retrieval method using 10mM sodium citrate (pH 6.0)
and microwaved for 8-15 min before commencing with IHC staining protocol.
Target
Function
Spectrin is the major constituent of the cytoskeletal network underlying the erythrocyte plasma
membrane. It associates with band 4.1 and actin to form the cytoskeletal superstructure of the
erythrocyte plasma membrane.
Involvement in disease
Defects in SPTB are the cause of elliptocytosis type 3 (EL3) [MIM:182870]. EL3 is a Rhesusunlinked form of hereditary elliptocytosis, a genetically heterogeneous, autosomal dominant
hematologic disorder. It is characterized by variable hemolytic anemia and elliptical or oval red
cell shape.
Defects in SPTB are the cause of spherocytosis type 2 (SPH2) [MIM:182870]; also known as
hereditary spherocytosis type 2 (HS2). Spherocytosis is a hematologic disorder leading to
chronic hemolytic anemia and characterized by numerous abnormally shaped erythrocytes which
are generally spheroidal. SPH2 is characterized by severe hemolytic anemia. Inheritance is
autosomal dominant.
Sequence similarities
Belongs to the spectrin family.
Contains 2 CH (calponin-homology) domains.
Contains 17 spectrin repeats.
Post-translational
modifications
The first phosphorylation event occurs on Ser-2114, followed by Ser-2125, Ser-2123, Ser-2128,
Ser-2117, and Thr-2110.
Cellular localization
Cytoplasm > cytoskeleton. Cytoplasm > cell cortex.
Anti-beta 1 Spectrin antibody [4C3] images
2
Immunocytochemistry/Immunofluorescence
analysis of beta 1 Spectrin (green) showing
staining in the cytoplasm of A431 cells (right)
compared to a negative control without
primary antibody (left). Formalin-fixed cells
were permeabilized with 0.1% Triton X-100 in
Immunocytochemistry/ Immunofluorescence -
TBS for 5-10 minutes and blocked with 3%
Anti-beta 1 Spectrin [4C3] antibody (ab2808)
BSA-PBS for 30 minutes at room
temperature. Cells were incubated with
ab2808 in 3% BSA-PBS at a dilution of 1:20
and incubated overnight at 4 ºC in a
humidified chamber. Cells were washed with
PBST and incubated with a DyLightconjugated secondary antibody in PBS at
room temperature in the dark. F-actin (red)
was stained with a fluorescent red phalloidin
and nuclei (blue) were stained with Hoechst or
DAPI. Images were taken at a magnification
of 60x.
Immunocytochemistry/Immunofluorescence
analysis of beta 1 Spectrin (green) showing
staining in the cytoplasm of Hela cells (right)
compared to a negative control without
primary antibody (left). Formalin-fixed cells
were permeabilized with 0.1% Triton X-100 in
Immunocytochemistry/ Immunofluorescence -
TBS for 5-10 minutes and blocked with 3%
Anti-beta 1 Spectrin [4C3] antibody (ab2808)
BSA-PBS for 30 minutes at room
temperature. Cells were incubated with
ab2808 in 3% BSA-PBS at a dilution of 1:100
and incubated overnight at 4 ºC in a
humidified chamber. Cells were washed with
PBST and incubated with a DyLightconjugated secondary antibody in PBS at
room temperature in the dark. F-actin (red)
was stained with a fluorescent red phalloidin
and nuclei (blue) were stained with Hoechst or
DAPI. Images were taken at a magnification
of 60x.
3
Anti-beta 1 Spectrin antibody [4C3] (ab2808)
at 1/1000 dilution + HeLa cell lysate at 25 µg
Western blot - Anti-beta 1 Spectrin [4C3] antibody
(ab2808)
ab2808 labelling beta 1 Spectrin in the
cytoplasm of Human prostate
carcinoma (right) compared with a negative
control (left) by Immunohistochemistry
(formalin-PFA-fixed paraffin-embedded
sections). To expose target proteins, antigen
Immunohistochemistry (Paraffin-embedded
retrieval method was performed using 10mM
sections) - Anti-beta 1 Spectrin antibody [4C3]
sodium citrate (pH 6.0) microwaved for 8-15
(ab2808)
min. Following antigen retrieval, tissues were
blocked in 3% H2O2-methanol for 15 min at
room temperature. Tissue sections were
incubated with the primary antibody (1:20 in
3% BSA-PBS) overnight at 4°C. A HRPconjugated anti-mouse was used as the
secondary antibody, followed by colorimetric
detection using a DAB kit. Tissues were
counterstained with hematoxylin and
dehydrated with ethanol and xylene to prep for
mounting.
4
ab2808 labelling beta 1 Spectrin in the
membrane of Human skeletal muscle (right)
compared with a negative control (left) by
Immunohistochemistry (formalin-PFA-fixed
paraffin-embedded sections). To expose
target proteins, antigen retrieval method was
Immunohistochemistry (Paraffin-embedded
performed using 10mM sodium citrate (pH
sections) - Anti-beta 1 Spectrin antibody [4C3]
6.0) microwaved for 8-15 min. Following
(ab2808)
antigen retrieval, tissues were blocked in 3%
H2O2-methanol for 15 min at room
temperature. Tissue sections were incubated
with the primary antibody (1:200 in 3% BSAPBS) overnight at 4°C. A HRP-conjugated
anti-mouse was used as the secondary
antibody, followed by colorimetric detection
using a DAB kit. Tissues were counterstained
with hematoxylin and dehydrated with ethanol
and xylene to prep for mounting.
ab2808 labelling beta 1 Spectrin in the
cytoplasm and membrane of Mouse skeletal
muscle (right) compared with a negative
control (left) by Immunohistochemistry
(formalin-PFA-fixed paraffin-embedded
sections). To expose target proteins, antigen
Immunohistochemistry (Paraffin-embedded
retrieval method was performed using 10mM
sections) - Anti-beta 1 Spectrin antibody [4C3]
sodium citrate (pH 6.0) microwaved for 8-15
(ab2808)
min. Following antigen retrieval, tissues were
blocked in 3% H2O2-methanol for 15 min at
room temperature. Tissue sections
were incubated with the primary antibody
(1:20 in 3% BSA-PBS) overnight at 4°C. A
HRP-conjugated anti-mouse was used as
the secondary antibody, followed by
colorimetric detection using a DAB kit.
Tissues were counterstained with hematoxylin
and dehydrated with ethanol and xylene to
prep for mounting.
5
IHC image of ab2808 staining in human
skeletal muscle formalin fixed paraffin
embedded tissue section, performed on a
Leica BondTM 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 ab2808, 1/200 dilution, for 15
mins at room temperature and detected using
Immunohistochemistry (Formalin/PFA-fixed
paraffin-embedded sections) - Anti-beta 1 Spectrin
antibody [4C3] (ab2808)
an HRP conjugated compact polymer system.
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.
Flow cytometry analysis of beta 1 Spectrin
showing positive staining in the cytoplasm of
SH-SY5Y cells compared to an isotype
control (blue). Cells were harvested, adjusted
to a concentration of 1-5x10^6 cells/ml, fixed
with 2% paraformaldehyde and washed with
PBS. Cells were penetrated by dropping the
supernatant, adding 90% methanol and
incubated for 10 minutes at room
temperature. Follwing penetration, cells were
blocked with a 2% solution of BSA-PBS for
Flow Cytometry - Anti-beta 1 Spectrin antibody
30 min at room temperature and incubated
[4C3] (ab2808)
with ab2808 (1 ug/test) for 60 min at room
temperature. Cells were then incubated for 40
min at room temperature in the dark using a
Dylight 488-conjugated goat anti-mouse IgG
(H+L) secondary antibody and re-suspended
in PBS for FACS analysis.
6
Flow cytometry analysis of beta 1 Spectrin
showing positive staining in the cytoplasm of
NIH/3T3 cells compared to an isotype control
(blue). Cells were harvested, adjusted to a
concentration of 1-5x10^6 cells/ml, fixed with
2% paraformaldehyde and washed with PBS.
Cells were penetrated by dropping the
supernatant, adding 90% methanol and
incubated for 10 minutes at room
temperature. Follwing penetration, cells were
blocked with a 2% solution of BSA-PBS for
Flow Cytometry - Anti-beta 1 Spectrin antibody
30 min at room temperature and incubated
[4C3] (ab2808)
with ab2808 (0.5 ug/test) for 60 min at room
temperature. Cells were then incubated for 40
min at room temperature in the dark using a
Dylight 488-conjugated goat anti-mouse IgG
(H+L) secondary antibody and re-suspended
in PBS for FACS analysis.
Flow cytometry analysis of beta 1
Spectrin showing positive staining in the
cytoplasm of Hela cells compared to an
isotype control (blue). Cells were harvested,
adjusted to a concentration of 1-5x10^6
cells/ml, fixed with 2% paraformaldehyde and
washed with PBS. Cells were penetrated by
dropping the supernatant, adding 90%
methanol and incubated for 10 minutes at
room temperature. Follwing penetration, cells
were blocked with a 2% solution of BSA-PBS
Flow Cytometry - Anti-beta 1 Spectrin antibody
for 30 min at room temperature and incubated
[4C3] (ab2808)
with ab2808 (1 ug/test) for 60 min at room
temperature. Cells were then incubated for 40
min at room temperature in the dark using a
Dylight 488-conjugated goat anti-mouse IgG
(H+L) secondary antibody and re-suspended
in PBS for FACS analysis.
7
Overlay histogram showing SH-SY5Y cells
stained with ab2808 (red line). The cells were
fixed with 4% paraformaldehyde (10 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 (ab2808,
Flow Cytometry-Anti-beta 1 Spectrin antibody
[4C3](ab2808)
1/50 dilution) for 30 min at 22ºC. The
secondary antibody used was DyLight® 488
goat anti-mouse 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.
Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"
Our Abpromise to you: Quality guaranteed and expert technical support
Replacement or refund for products not performing as stated on the datasheet
Valid for 12 months from date of delivery
Response to your inquiry within 24 hours
We provide support in Chinese, English, French, German, Japanese and Spanish
Extensive multi-media technical resources to help you
We investigate all quality concerns to ensure our products perform to the highest standards
If the product does not perform as described on this datasheet, we will offer a refund or replacement. For full details of the Abpromise,
please visit http://www.abcam.com/abpromise or contact our technical team.
Terms and conditions
Guarantee only valid for products bought direct from Abcam or one of our authorized distributors
8