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BCAR1 / Breast cancer anti-estrogen resistance protein 1 · Western blot design guide
Design a Western Blot for BCAR1
Source-linked BCAR1 Western blot protocol options, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-BCAR1 WB antibodies. Everything you need to plan the experiment before you commit precious samples.
Evidence assembled September 2026 · For research use; verify linked source records and product datasheet before use
BCAR1 Western Blot Experimental Design Guide
Expected bands, source-linked protocol options, controls and antibodies — the at-a-glance facts below, then the full design guide.
Must know before running
| Expected band | ~93.4 kDa | |
| Gel | 8–10% (standard starting point) | |
| Positive control | Bronchus (IHC candidate; verify WB) +4 more | |
| Negative control | Bone marrow (IHC candidate; verify WB) |
Important caveats
Reasons your observed band may differ from the expected size.
| PTM | Phosphorylated + Acetylated | |
| Caveat | Phosphorylation-state controls | |
| Gene-set association | MSigDB Hallmark membership | |
| Isoform | 8 isoform(s) |
Section 1
Source-Linked BCAR1 Western Blot Protocol Options
The M00960-2 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | A431 cell lysate (catalog M00960-2) |
| Gel % | 8–10% (standard starting point) |
| Load | 20–30 µg total protein per lane; optimize for abundance (standard starting point) |
| Transfer | Standard semi-dry transfer; verify efficiency (standard starting point) |
| Membrane | 0.45 µm PVDF (standard starting point) |
| Blocking | 5% milk or 5% BSA in TBST (standard starting point) |
| Primary antibody | M00960-2; use the WB datasheet starting dilution (standard starting point) |
| Primary incubation | Overnight at 4 °C (standard starting point) |
| Secondary antibody | Species-matched HRP conjugate at validated dilution (standard starting point) |
| Secondary incubation | 1 h at room temperature (standard starting point) |
| Wash | 3 × 5 min in TBST (standard starting point) |
| Detection | ECL; bracket exposures to avoid saturation (standard starting point) |
Section 2
What Is the Expected BCAR1 Western Blot Band Size?
BCAR1 is predicted at 93.4 kDa; isoforms and phosphorylation could affect migration, but no empirical band size or visible effect is established.
| Band near 93.4 kDa | Consistent with the predicted BCAR1 mass; confirm identity with antibody controls. |
| Band below 93.4 kDa | Could represent an alternatively spliced BCAR1 isoform; its migration is not established. |
| Band above 93.4 kDa | Could reflect altered migration associated with phosphorylation; the cause requires testing. |
| Several bands at different positions | Could reflect BCAR1 isoforms 1 through 8 or different phosphorylation states; distinct bands are not established. |
💡Expected BCAR1 appearanceBCAR1 has a predicted mass of 93.4 kDa, but no empirical band size is supplied; use antibody specificity controls to identify any band and test possible effects of splicing or phosphorylation.
| UniProt predicted mass | Places the reference BCAR1 sequence at 93.4 kDa. |
| Isoforms 1 through 8 | Alternative splicing could change apparent size; individual isoform sizes are not supplied. |
| Phosphotyrosine at residue 128 | Could affect migration, but a visible shift is not established. |
| Phosphoserine at residue 134 | Could affect migration, but a visible shift is not established. |
| Phosphothreonine at residue 269 | Could affect migration, but a visible shift is not established. |
| Situation | Likely cause | Next action |
|---|---|---|
| Band higher than expected | Phosphorylation is possible, but its effect on BCAR1 migration is unproven. | Compare phosphatase-treated and untreated lysates and confirm band identity. |
| Band lower than expected | An alternatively spliced isoform is possible; its size is unknown. | Check isoform expression and confirm the band with an independent BCAR1 antibody. |
| Broad smear instead of sharp band | Multiple phosphorylation states are possible, but the smear's cause is unknown. | Compare phosphatase-treated lysate and check sample quality. |
| Multiple bands | BCAR1 has eight named isoforms and documented phosphorylation sites; neither proves resolvable bands. | Compare isoform expression and phosphatase treatment, then verify band identity. |
| Weak or no signal | BCAR1 detection in the tested lysate is unconfirmed. | Check loading and transfer and test a validated BCAR1-positive lysate. |
Experiment evidence
Western blot images showing BCAR1 band
M00960-2 · BCAR1
M00960Y410 · BCAR1
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for BCAR1 Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for BCAR1 in Western blot, you can use bronchus lysate, which HPA scores High.
Positive control: Bronchus (IHC candidate; verify WB)
Negative control: Bone marrow (IHC candidate; verify WB)
Loading controls: Run GAPDH, β-actin, and a total-protein stain alongside the samples.
⚠️Feasibility: BCAR1 is intracellular, and HPA identifies bone marrow as a not-detected comparator.
HPA tissue expression evidence for BCAR1
Comprehensive Human Protein Atlas IHC scoring per tissue. Rows are taken directly from the HPA tissue chart — click any row's HPA link to view the source.
Higher expression tissues · candidate positive controls from IHC
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Bronchus | basal cells | High | Protein (IHC) | HPA → |
| Cerebellum | Purkinje cells | High | Protein (IHC) | HPA → |
| Fallopian tube | ciliated cells (ciliary rootlets) | High | Protein (IHC) | HPA → |
| Gallbladder | glandular cells | High | Protein (IHC) | HPA → |
| Rectum | glandular cells | High | Protein (IHC) | HPA → |
Lower expression tissues · IHC evidence, not confirmed WB-negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Bone marrow | hematopoietic cells | Not detected | Protein (IHC) | HPA → |
| Caudate | glial cells | Not detected | Protein (IHC) | HPA → |
| Heart muscle | cardiomyocytes | Not detected | Protein (IHC) | HPA → |
| Parathyroid gland | glandular cells | Not detected | Protein (IHC) | HPA → |
| Prostate | glandular cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced BCAR1 Western Blot Tips
Deeper troubleshooting and optimisation questions for BCAR1, answered from its protein features.
How should BCAR1 band migration be interpreted?
Band shift · Use the separately labelled calculated mass and catalog-observed evidence above. A sequence annotation does not establish an observed migration shift. Verify target identity with orthogonal controls.
How might BCAR1 isoforms affect band interpretation?
Isoforms · UniProt lists eight isoforms. Isoform 4 lacks canonical residues 64–211; isoforms 2 and 5–8 have different N termini, and isoform 3 changes residue 304. Check which isoforms the antibody recognizes before assigning bands. The supplied features do not establish which isoforms are expressed in your sample.
Choose an epitope shared by the isoforms of interest and verify it against their sequences. N-terminal residues 1–4 vary in isoforms 2 and 4–8, isoform 4 lacks residues 64–211, and isoform 3 changes residue 304. An antibody to an altered or absent region may miss an isoform.
Choose an epitope shared by the isoforms of interest and verify it against their sequences. N-terminal residues 1–4 vary in isoforms 2 and 4–8, isoform 4 lacks residues 64–211, and isoform 3 changes residue 304. An antibody to an altered or absent region may miss an isoform.
Which BCAR1 sites support phospho-specific detection?
PTM · UniProt lists phosphotyrosines at 128 (by SRC), 234, 249 (by ABL1), 362, 372 and 410; phosphoserines at 134, 139, 292, 428, 437 and 639; and phosphothreonine at 269. These are UniProt canonical coordinates, which may differ from antibody or paper numbering. Match the antibody’s stated site to the intended sequence.
Measure a specified phosphosite and total BCAR1 in matched samples, then compare the phosphosite signal relative to total BCAR1. Record the antibody’s site-numbering convention: UniProt coordinates include SRC-modified Tyr128 and ABL1-modified Tyr249. The listed sites do not establish that phosphorylation changes under a particular treatment.
Measure a specified phosphosite and total BCAR1 in matched samples, then compare the phosphosite signal relative to total BCAR1. Record the antibody’s site-numbering convention: UniProt coordinates include SRC-modified Tyr128 and ABL1-modified Tyr249. The listed sites do not establish that phosphorylation changes under a particular treatment.
Does this guide establish induction of BCAR1?
Induction · No general induction response is established by this guide. A pathway or gene-set association is not evidence of induction in a particular specimen. Verify the relevant treatment and control in a target-specific experiment.
How should transfer be checked for BCAR1?
Transfer · Standard workflow guidance: verify transfer efficiency for the intended target size before interpreting a weak signal. Use total-protein assessment and optimize transfer for the membrane, gel and apparatus; the labelled catalog values take precedence.
How should blocking be optimized?
Blocking · Standard workflow guidance: follow the M00960-2 datasheet where specified. Otherwise compare 5% milk or 5% BSA in TBST; for a phospho-specific assay start with BSA. Optimize background and specific signal with matched controls.
How should BCAR1 be quantified?
Quantitation · Standard workflow guidance: quantify only a validated target band within the linear exposure range. Use consistent sample preparation and loading, retain biological replicates, and avoid interpreting saturation or loading differences as regulation.
Why might BCAR1 differ from its predicted 93.4 kDa?
Interpretation · BCAR1 has eight isoforms and multiple modified residues, but these features alone do not establish a visible shift or explain any difference from 93.4 kDa. No empirical apparent band size was supplied. Compare the observed band with the predicted size and evaluate isoform and phosphorylation signals separately.
UniProt reports BCAR1 at focal adhesions following integrin engagement and describes the unphosphorylated form as cytoplasmic by similarity. Keep engagement conditions and sampling consistent when comparing cells. A change in localization alone does not establish a change in total BCAR1 abundance or phosphorylation.
Check antibody epitope coverage against the eight isoforms, especially isoform 4’s deletion of residues 64–211 and the N-terminal variants. Compare total BCAR1 and site-specific signals where relevant. BCAR1 also has listed phosphorylation sites, but their presence alone cannot identify an unexpected band or prove a mobility shift.
UniProt reports BCAR1 at focal adhesions following integrin engagement and describes the unphosphorylated form as cytoplasmic by similarity. Keep engagement conditions and sampling consistent when comparing cells. A change in localization alone does not establish a change in total BCAR1 abundance or phosphorylation.
Check antibody epitope coverage against the eight isoforms, especially isoform 4’s deletion of residues 64–211 and the N-terminal variants. Compare total BCAR1 and site-specific signals where relevant. BCAR1 also has listed phosphorylation sites, but their presence alone cannot identify an unexpected band or prove a mobility shift.
Boster reagents
BCAR1 Western Blot Antibodies
Catalog antibodies with Western blot application and product-specific WB images. Evaluate suitability with the reported sample, controls and experimental conditions.
Real WB data
Anti-BCAR1 Rabbit Monoclonal Antibody
Cat # M00960-2
Real WB data
Anti-Phospho-BCAR1 (Y410) Rabbit Monoclonal Antibody
Cat # M00960Y410
Two the supplier rabbit monoclonal antibodies are listed: M00960-2 for BCAR1 and M00960Y410 for phospho-BCAR1 (Y410). Both have WB images, showing A431 lysate and pervanadate-treated HeLa lysate, respectively. These examples establish tested contexts, not broader validation.
Which to pick: Choose M00960-2 for total BCAR1; its listed reactivity is human, mouse and rat. Choose M00960Y410 for Y410-phosphorylated BCAR1 in human samples. Its WB image uses pervanadate-treated HeLa lysate; the M00960-2 image uses A431 lysate.
Source: BosterBio BCAR1 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry P56945.
- Human Protein Atlas. BCAR1 tissue expression.
- PMC8702347 — target-verified WB comparison