PRKDC / DNA-dependent protein kinase catalytic subunit · Western blot design guide

Design a Western Blot for PRKDC

Real validated PRKDC Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-PRKDC 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

Western blot protocol sheet for PRKDC: expected band ~469.1 kDa, hero antibody PA1970, catalog values and labelled standard workflow; separate PMC comparisons on the guide — Printable **PRKDC** Western blot protocol sheet — expected band ~469.1 kDa, antibody PA1970, controls and PMC citations. Open the full PRKDC WB guide →

Navigate to: Protocols Band Controls Antibodies

PRKDC Western Blot Experimental Design Guide

Expected bands, validated protocols, controls and antibodies — the at-a-glance facts below, then the full design guide.

Must know before running

	Expected band	~469.1 kDa
	Observed band	~469 kDa
	Gel	8% (catalog PA1970)
	Positive control ⓘ	Adipose tissue (IHC candidate; verify WB) +4 more
	Negative control ⓘ	Suggested KO / knockdown lysate

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	2 isoform(s)

Section 1

Real Curated PRKDC Western Blot Protocols

The PA1970 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.

Recommended Western blot protocol parameters
Sample / lysate	human SiHa (catalog PA1970)
Gel %	8% (catalog PA1970)
Load	30 ug; reducing conditions (catalog PA1970)
Transfer	a nitrocellulose membrane at 150 mA for 50-90 minutes (catalog PA1970)
Membrane	nitrocellulose membrane (catalog PA1970)
Blocking	5% non-fat milk/TBS for 1.5 hour at RT (catalog PA1970)
Primary antibody	PA1970 · 0.5 μg/mL (catalog PA1970)
Primary incubation	overnight at 4°C (catalog PA1970)
Secondary antibody	goat anti-rabbit IgG-HRP, 1:5000 (catalog PA1970)
Secondary incubation	1.5 hour at RT (catalog PA1970)
Wash	TBS-0.1%Tween 3 times with 5 minutes each (catalog PA1970)
Detection	ECL (catalog PA1970)

Section 2

What Is the Expected PRKDC Western Blot Band Size?

PRKDC is predicted at 469.1 kDa and observed at ~469 kDa; these agree within rounding, while distinct isoform migration is unestablished.

What am I looking at on my blot?
Band at ~469 kDa	Matches the reported PRKDC band and its 469.1 kDa predicted mass.
Additional band at an unspecified size	Could represent isoform 1 or 2 if the isoforms differ in mass and resolve.
Close doublet near ~469 kDa	Could reflect different phosphorylation states; a mobility difference is not established.
Band at ~469 kDa in a nuclear fraction	Consistent with PRKDC localization to the nucleus.

💡Expected PRKDC appearanceExpect a band at the empirically observed ~469 kDa, close to the predicted 469.1 kDa; confirm its identity with appropriate antibody and sample controls, especially if additional bands appear.

How each factor affects band size
Predicted PRKDC mass	469.1 kDa predicts a band near the observed ~469 kDa.
Splice isoform 1	Its individual mass and mobility relative to isoform 2 are not supplied.
Splice isoform 2	Its individual mass and mobility relative to isoform 1 are not supplied.
Alternative splicing of isoforms 1 and 2	Could yield different band sizes if the isoforms differ in mass and resolve.

Why is my band missing or off?
Situation	Likely cause	Next action
No band in lysate	Nuclear PRKDC may be poorly recovered during extraction.	Check nuclear extraction and include a positive control lysate.
Band higher than expected	The cause is not established by the supplied features.	Verify size markers and band identity with an independent antibody.
Band lower than expected	The large PRKDC protein may have degraded during sample preparation.	Prepare fresh lysate with protease inhibitors and compare with a positive control.
Broad smear instead of sharp band	Degradation or uneven transfer of this large protein may blur the signal.	Check sample integrity and optimize transfer for high-mass proteins.
Multiple bands	Isoforms 1 and 2 or nonspecific antibody binding are possible; distinct isoform migration is unestablished.	Compare with an independent antibody and a PRKDC-depleted control.
Weak or no signal	Recovery or transfer of the large nuclear protein may be insufficient.	Check nuclear extraction, transfer, and a positive control.

Source: UniProt P78527 Human Protein Atlas product datasheet (PA1970)

Experiment evidence

Western blot images showing PRKDC band

PA1970 · PRKDC

M00645 · PRKDC

P00645 · PRKDC

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Real WB data behind the band-size expectations in this section.

Sample controls for PRKDC Western blot

🧪HPA-IHC candidate guidance (verify in WB): For positive controls for PRKDC in Western blot, you can use adipose tissue, which has high HPA expression.

Positive control: Adipose tissue (IHC candidate; verify WB)

Negative control: Suggested KO / knockdown lysate

Loading controls: Run GAPDH, β-actin, and a total-protein stain alongside the samples.

⚠️Feasibility: No HPA tissue is undetected, so use siRNA knockdown or a KO line as the negative control.

HPA tissue expression evidence for PRKDC

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
Adipose tissue	adipocytes	High	Protein (IHC)	HPA →
Adrenal gland	glandular cells	High	Protein (IHC)	HPA →
Appendix	glandular cells	High	Protein (IHC)	HPA →
Bone marrow	hematopoietic cells	High	Protein (IHC)	HPA →
Breast	glandular cells	High	Protein (IHC)	HPA →

Lower expression tissues · IHC evidence, not confirmed WB-negative controls

Tissue	Cell type	Level	Evidence	Source
Liver	cholangiocytes	Medium	Protein (IHC)	HPA →
Smooth muscle	smooth muscle cells	Medium	Protein (IHC)	HPA →
Adipose tissue	adipocytes	High	Protein (IHC)	HPA →
Adrenal gland	glandular cells	High	Protein (IHC)	HPA →
Appendix	glandular cells	High	Protein (IHC)	HPA →

Section 3

Advanced PRKDC Western Blot Tips

Deeper troubleshooting and optimisation questions for PRKDC, answered from its protein features.

How should PRKDC 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.

Could PRKDC isoforms produce different bands?

Isoforms · UniProt lists two isoforms. Isoform 2 lacks residues 3799–3829 relative to isoform 1, so it is shorter by 31 amino acids. Check whether the antibody epitope includes that interval before comparing isoform signals.

Which PRKDC phosphorylation sites matter for phospho-specific detection?

PTM · UniProt lists autophosphorylation at Ser2056, Thr2609, Ser2612, Thr2638 and Thr2647. Specify the site when selecting a phospho-specific antibody. These are UniProt coordinates; an antibody or paper may use a different numbering convention.

Does this guide establish induction of PRKDC?

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 PRKDC Western blot?

Transfer · PRKDC is predicted to be 469.1 kDa. Use a transfer setup validated for proteins of this size, and check the membrane and post-transfer gel for incomplete transfer before interpreting a weak band.

How should blocking be optimized?

Blocking · Standard workflow guidance: follow the PA1970 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 PRKDC abundance be quantified across fractions?

Quantitation · PRKDC is listed in the nucleus, nucleolus and cytosol. Compare like fractions across samples and normalize within each fraction; a change in one fraction alone does not establish a change in total PRKDC abundance.

Why might PRKDC migrate near 469 kDa?

Interpretation · UniProt predicts 469.1 kDa, close to the supplied observed band of about 469 kDa. Its listed phosphorylation and acetylation sites do not, by themselves, establish a visible mobility shift.

First compare bands with the predicted 469.1 kDa protein and the 31-residue deletion in isoform 2. The listed modifications alone cannot identify an unexpected band or establish a visible shift. Check whether the antibody epitope spans residues 3799–3829; UniProt lists phospho-Ser3821 within that deleted interval.

Boster reagents

PRKDC 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

Western blot analysis of PRKDC using anti-PRKDC antibody (PA1970). Electrophoresis was performed on a 8% SDS-PAGE gel at 80V (Stacking gel) / 120V (Resolving gel) for 2 hours. The sample well of each lane was loaded with 30 ug of sample under reducing conditions. Lane 1: human SiHa whole cell lysates. After electrophoresis, proteins were transferred to a nitrocellulose membrane at 150 mA for 50-90 minutes. Blocked the membrane with 5% non-fat milk/TBS for 1.5 hour at RT. The membrane was incubated with rabbit anti-PRKDC antigen affinity purified polyclonal antibody (PA1970) at 0.5 μg/mL overnight at 4°C, then washed with TBS-0.1%Tween 3 times with 5 minutes each and probed with a goat anti-rabbit IgG-HRP secondary antibody (Catalog # BA1054) at a dilution of 1:5000 for 1.5 hour at RT. The signal is developed using an ECL Plus Western Blotting Substrate (Catalog # AR1196-200) with Tanon 5200 system. A specific band was detected for PRKDC at approximately 469 kDa. The expected band size for PRKDC is at 469 kDa.

Anti-DNA PKcs/PRKDC Antibody Picoband®

Cat # PA1970

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Real WB data Western blot analysis of DNA-PKcs expression in HeLa cell lysate.

Anti-DNA-PKcs PRKDC Rabbit Monoclonal Antibody

Cat # M00645

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Real WB data Western blot analysis of Phospho-DNA PKcs (Ser2056) expression in alkaline treated Jurkat cell lysate.

Anti-Phospho-DNA PKcs (S2056) PRKDC Rabbit Monoclonal Antibody

Cat # P00645

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Three listed antibodies have WB images: PA1970 in human SiHa lysate with a band near 469 kDa, M00645 in HeLa lysate, and phospho-Ser2056 P00645 in alkaline-treated Jurkat lysate. These examples show tested contexts, not validation across every listed species.

Which to pick: For total PRKDC, choose PA1970 if the documented SiHa band and protocol are useful, or M00645 for its HeLa WB example; both list human, mouse, and rat reactivity. For Ser2056 phosphorylation, choose P00645, which lists human reactivity only.

Source: BosterBio PRKDC gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.

References

UniProt Consortium. UniProt entry P78527.
Human Protein Atlas. PRKDC tissue expression.
PMC4320452 — target-verified WB comparison
PMC6825841 — target-verified WB comparison
PMC9220184 — target-verified WB comparison