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- Table of Contents
CSN2 / Beta-casein · Western blot design guide
Design a Western Blot for CSN2
Source-linked CSN2 Western blot protocol options, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-CSN2 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
CSN2 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 | ~25.4 kDa | |
| Gel | 12–15% (standard starting point) | |
| Positive control | Lactating breast (IHC candidate; verify WB) | |
| Negative control | Adipose tissue (IHC candidate; verify WB) |
Important caveats
Reasons your observed band may differ from the expected size.
| PTM | Phosphorylated + Cleaved | |
| Caveat | Phosphorylation-state controls | |
| Gene-set association | MSigDB C7 membership | |
| Isoform | 1 isoform(s) |
Section 1
Source-Linked CSN2 Western Blot Protocol Options
The M03701 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | Human milk lysate (catalog M03701) |
| Gel % | 12–15% (standard starting point) |
| Load | 20–30 µg total protein per lane; optimize for abundance (standard starting point) |
| Transfer | Short semi-dry transfer; verify retention (standard starting point) |
| Membrane | 0.45 µm PVDF (standard starting point) |
| Blocking | 5% milk or 5% BSA in TBST (standard starting point) |
| Primary antibody | M03701; 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 CSN2 Western Blot Band Size?
Beta-casein has a predicted 25.4 kDa precursor; signal-peptide cleavage and phosphorylation may affect migration, but no empirical band size is supplied.
| Band near 25.4 kDa | Consistent with the predicted beta-casein precursor; confirm identity with controls |
| Band below 25.4 kDa | Could reflect removal of the 1–15 signal peptide; mature migration is not supplied |
| Little or no band in whole-cell lysate | Consistent with secretion of beta-casein |
| Closely spaced bands near the expected region | Could reflect the annotated phosphorylation forms; distinct migration is not established |
💡Expected CSN2 appearanceThe UniProt precursor mass is 25.4 kDa; signal-peptide removal and phosphorylation may affect migration, but no empirical band size is supplied, so confirm any candidate band with identity controls.
| UniProt predicted precursor mass | 25.4 kDa is the sequence-based starting estimate |
| Signal peptide at residues 1–15 | Cleavage produces a smaller mature protein; its apparent band size is not supplied |
| Phosphothreonine at residue 18 in form 5-P | May affect apparent migration; no visible shift is demonstrated |
| Phosphoserine at residue 24 in forms 1-P through 5-P | Phosphorylation state may affect apparent migration; no distinct band sizes are supplied |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | Beta-casein is secreted | Check a secreted-protein sample and include a positive control |
| Band higher than expected | The cause is not established; phosphorylation may affect migration | Compare with a positive control and verify antibody specificity |
| Band lower than expected | Signal-peptide removal may reduce the protein's mass | Compare precursor and mature sequence coverage if available, and verify band identity |
| Multiple bands | Annotated phosphorylation forms could differ in migration, but separation is unproven | Check antibody specificity and compare bands with a positive control |
| Weak or no signal | A whole-cell sample may contain little secreted beta-casein | Test a secreted-protein sample and confirm antibody performance with a positive control |
Experiment evidence
Western blot images showing CSN2 band
M03701 · CSN2
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for CSN2 Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for CSN2 in Western blot, you can use lactating breast tissue, where HPA reports high expression.
Positive control: Lactating breast (IHC candidate; verify WB)
Negative control: Adipose tissue (IHC candidate; verify WB)
Loading controls: Run GAPDH, β-actin, and a total-protein stain such as Ponceau alongside the samples.
⚠️Feasibility: CSN2 is secreted, so conditioned medium may give a stronger signal than whole-cell lysate.
HPA tissue expression evidence for CSN2
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 |
|---|---|---|---|---|
| Lactating breast | ductal cells | High | Protein (IHC) | HPA → |
Lower expression tissues · IHC evidence, not confirmed WB-negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | adipocytes | Not detected | Protein (IHC) | HPA → |
| Adrenal gland | glandular cells | Not detected | Protein (IHC) | HPA → |
| Appendix | glandular cells | Not detected | Protein (IHC) | HPA → |
| Bone marrow | hematopoietic cells | Not detected | Protein (IHC) | HPA → |
| Bronchus | respiratory epithelial cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced CSN2 Western Blot Tips
Deeper troubleshooting and optimisation questions for CSN2, answered from its protein features.
How does signal peptide processing affect the expected CSN2 band?
Band shift · UniProt lists a signal peptide at residues 1..15, so the secreted protein is expected to lack that segment. The supplied 25.4-kDa value describes the precursor; no measured mass for the processed protein is supplied.
Could annotated CSN2 isoforms explain multiple bands?
Isoforms · The supplied record lists one isoform and no alternative sequence. Do not assign additional bands to annotated isoforms on this evidence. Compare any unexpected bands with the signal peptide and phosphorylation features, then verify their identity experimentally.
Which CSN2 phosphorylation sites matter when comparing bands?
PTM · UniProt lists Thr18 in form 5-P; Ser21 in forms 4-P and 5-P; Ser23 in forms 3-P through 5-P; and Ser24 and Ser25 in forms 1-P through 5-P. These are UniProt precursor coordinates, which may differ from antibody or paper numbering. Phosphorylation alone does not establish a visible band shift.
Does this guide establish induction of CSN2?
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.
What transfer method to use for CSN2 Western blot?
Transfer · CSN2 is a 25.4-kDa precursor with a 1..15 signal peptide. Choose transfer conditions suitable for a protein around this size and verify retention using an appropriate marker. The supplied features do not specify one transfer method.
How should blocking be optimized?
Blocking · Standard workflow guidance: follow the M03701 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 CSN2 bands be quantified?
Quantitation · CSN2 is secreted and has several annotated phosphorylation forms. Compare equivalent secreted sample fractions, and state whether quantitation uses one band or all CSN2-reactive bands. The feature record does not establish that different forms resolve into separate bands.
Why might CSN2 migrate differently from its predicted mass?
Interpretation · The 25.4-kDa prediction is for the 226-residue precursor. CSN2 has a signal peptide at residues 1..15 and five listed phosphorylation sites. These features do not establish an apparent band size or explain a measured difference; no observed band is supplied.
Check whether its size is consistent with signal peptide removal or whether phosphorylation forms could be relevant. UniProt lists no glycosylation sites or alternative sequence here. None of these features identifies an unexpected band by itself; confirm its identity before assigning a cause.
Check whether its size is consistent with signal peptide removal or whether phosphorylation forms could be relevant. UniProt lists no glycosylation sites or alternative sequence here. None of these features identifies an unexpected band by itself; confirm its identity before assigning a cause.
Boster reagents
CSN2 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-beta Casein Rabbit Monoclonal Antibody
Cat # M03701
For CSN2 Western blots, the listed the supplier antibody is M03701, an anti-beta Casein rabbit monoclonal with stated human reactivity. Its Western blot image shows beta Casein expression in human milk lysate; the supplied evidence covers that tested context.
Which to pick: M03701 is the only listed option. Its Western blot image is from human milk lysate, making that the directly documented sample. Performance in other sample types or species is not documented here.
Source: BosterBio CSN2 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry P05814.
- Human Protein Atlas. CSN2 tissue expression.
- PMC5751224 — target-verified WB comparison
- PMC5064986 — target-verified WB comparison
- PMC11200946 — target-verified WB comparison