MHCD0400

antibody from Invitrogen Antibodies

Targeting: CD4

Provider product page for MHCD0400

Flow cytometry

Other assay

Antibody data

Antibody Data
Antigen structure
References [66]
Comments [0]
Validations
Other assay [10]

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Product number: MHCD0400 - Provider product page
Provider: Invitrogen Antibodies
Product name: CD4 Monoclonal Antibody (S3.5)
Antibody type: Monoclonal
Antigen: Other
Reactivity: Human
Host: Mouse
Isotype: IgG
Antibody clone number: S3.5
Vial size: 500 µL
Storage: 4° C, store in dark

CD4 protein structure - MHCD0400 shown in red.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: NULL

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: NULL

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: NULL

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: NULL

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: NULL

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: NULL

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 4 The BFNC R353G mutation blocks axonal enrichment of surface CD4-Q2C. (A) The R353G mutation reduced but did not abolish co-immunoprecipitation of CaM with CD4-Q2C from transfected HEK293T cells. beta-actin served as a loading control for total cell lysates (B) Surface immunostaining of WT or R353G mutant CD4-Q2C in hippocampal neurons (DIV 7-8). Camera lucida drawings (lower) of the inverted images of surface CD4-Q2C (upper) show the soma and dendrites (gray) and an axon (black). The axon was identified by the lack of MAP2 immunostaining in the neurons cotransfected with GFP. The R353G mutation blocked enrichment of CD4-Q2C on the axonal surface by increasing its somatodendritic surface expression. Arrows mark the main axon. Scale bars are 20 um. (C) In comparison to WT, the surface ""Axon/Dendrite"" fluorescence ratio of CD4-Q2C was reduced to 1 by the R353G mutation, whereas the surface ""AIS/Axon"" ratio was unaffected. (D) Background subtracted, mean intensity of surface CD4 fluorescence in the AIS, distal axons, soma, and major dendrites. The R353G mutation increased CD4-Q2C expression at the somatodendritic surface compared to WT. The sample number for each construct used in (C, D) was as follows: WT (n = 23), R353G (n = 18), and untransfected (n = 15). AU, arbitrary unit. Ave +- SEM (*p

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 2 Mutations in IQ motif abolish axonal enrichment of surface CD4-Q2C. (A) Schematic drawings (not to scale) of a human KCNQ2 subunit (accession #Y15065) and CD4-Q2C showing CaM-binding domain. The amino acids in bold text are critical residues in the CaM-binding consensus IQ motif in helix A. Mutations in the underlined amino acids have been shown to abolish (L339R, I340E, and A343D) or moderately decrease (R353G) CaM interaction with KCNQ2 [24] , [26] . Mutations in the amino acids colored red are associated with BFNC [43] . (B) Lysates from HEK293T cells expressing CaM and CD4-Q2C wild-type (WT) or mutant proteins (L339R, I340E, and A343D) were subjected to immunoprecipitation (IP) with the CD4 antibody. Immunoprecipitation and total cell lysates were analyzed by immunoblotting for CD4 and CaM. beta-actin served as a loading control. The L339R, I340E, and A343D mutations abolished co-immunoprecipitation of CaM with CD4-Q2C. (C) Surface immunostaining of CD4-Q2C WT or mutant proteins in hippocampal neurons (DIV 7-8). Camera lucida drawings (lower) of the inverted images of surface CD4-Q2C (upper) show the soma and dendrites (gray) and an axon (black). The axon was identified by immunostaining for the AIS marker phospho IkappaBalpha Ser32 (14D4) in the neurons cotransfected with GFP, which allows visualization of all neurites ( Figure S3 ). Arrows indicate the AIS. Scale bars are 20 um. The L339R, I340E, and A343D mutations abolished surface expression of CD4-Q2C at th

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 3 Mutations in IQ motif block CD4-Q2C expression in the axon. (A) Permeabilized immunostaining of CD4-Q2C wild-type (WT) or mutant (L339R, I340E, or A343D) in GFP-cotransfected hippocampal neurons (DIV 7-8). Camera lucida drawings (lower) of the inverted images of total CD4-Q2C (upper) show the soma and dendrites (gray) and an axon (black). Arrows indicate the AIS identified by phospho IkappaBalpha Ser32 (14D4) immunostaining. The L339R, I340E, and A343D mutations abolished total (surface and intracellular) expression of CD4-Q2C in the axon. (B) Background subtracted, mean intensity of total CD4 fluorescence in the AIS, distal axons, soma, and major dendrites. AU, arbitrary unit. The sample number for each construct used was as follows: WT (n = 18), L339R (n = 20), I340E (n = 12), A343D (n = 25), and untransfected (n = 20). Ave +- SEM (*p

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 6 CD4-Q2C accumulates in the ER after BFA treatment. (A) Schematic drawings of a hippocampal neuron containing a continuous network of the ER from the soma to the dendrites. Treatment with brefeldin-A (BFA) leads to inhibition of anterograde transport from the ER to the Golgi complex [44] . (B-C) The cultured hippocampal neurons (DIV 5) were treated with vehicle control (No BFA) or BFA (0.75 ug/ml) at 30 min post transfection with GFP and CD4-Q2C wild-type (WT) (B), I340E mutant (C), or R353G mutant (D). At 16 hr post-BFA treatment, permeabilized immunostaining was performed to visualize total (surface and intracellular) expression of CD4-Q2C (inverted images, upper). BFA treatment caused newly synthesized wild-type and all mutant proteins to accumulate at perinuclear regions in the soma and proximal dendrites. Arrows mark the main axon identified by the lack of MAP2 immunostaining in the neurons cotransfected with GFP. Camera lucida drawings (lower) show the soma and dendrites (gray) and an axon (black). Scale bars are 20 um.