High sensitivity & low background detection: Digoxigenin-labeled DNA & RNA probes

Digoxigenin-labeled DNA and RNA probes are the ideal choice when high sensitivity and low background detection are required e.g. for in situ hybridization experiments:

Higher sensitivity compared to fluorescence labeling: Signal amplification effect through antibody-based two-step detection^[1-5].
Lower background compared to Biotinylated probes: Digoxigenin antibody does not interact with endogenous biotin in biological samples^[1-5].

Such Digoxigenin-labeled DNA and RNA probes are conveniently prepared by enzymatic incorporation of Digoxigenin-labeled nucleotides (Tab. 1, Fig. 1). Detection is performed with Digoxigenin antibodies conjugated to a reporter enzyme (colorimetric or chemiluminescent read out) or a fluorescent dye thus offering flexible detection formats for various read-out requirements.

Figure 1: Comparable labeling efficiency of HighYield T7 Digoxigenin RNA Labeling Kit (JBS) compared to DIG RNA Labeling Mix (Roche).
Digoxigenin (DIGX)-labeled RNA was synthesized by in vitro transcription for 30 min at 37°C according to the HighYield T7 Digoxigenin RNA Labeling Kit (DIGX-RNA (JBS)) or using HighYield T7 Digoxigenin RNA Labeling Kit but nucleotide components exchanged by DIG RNA Labeling Mix (Roche, #11277073910) (DIGX-RNA (Roche)).
Silica-membrane purified RNA (1423 nt) was subsequently spotted on a nylon membrane and detected with Anti-Digoxigenin-AP-Conjugate (Roche, #11093274910, 1:5000 dilution).
+: positive control / 3’-Digoxigenin-labeled Control Oligonucleotide (60 nt)
-: negative control / Unlabeled purified RNA w/o Digoxigenin-11-UTP during in vitro transcription
DIGX-RNA (JBS): Purified Digoxigenin-labeled RNA synthesized with HighYield T7 Digoxigenin RNA Labeling Kit
DIGX-RNA (Roche): Purified Digoxigenin-labeled RNA synthesized using HighYield T7 Digoxigenin RNA Labeling Kit but nucleotide components exchanged by DIG RNA Labeling Mix (Roche, #11277073910)

Non-radioactive Labeling of DNA and RNA

Table 1: Overview of available products for Digoxigenin Labeling

Product	Cat. No.	Application
Single Nucleotides
DIG-11-UTP	NU-821-DIGX	In vitro Transcription- based random labeling of RNA probes
DIG-11-dUTP	NU-803-DIGX	PCR-based random labeling of DNA probes
DIG-11-dCTP	NU-809-DIGX	PCR-based random labeling of DNA probes
DIG-11-ddUTP	NU-1619-DIGX	3‘-End oligonucleotide labeling with Terminal deoxynucleotidyl Transferase (TdT)
DIG-11-ddCTP	NU-850-DIGX	Potential substrates of Terminal deoxynucleotidyl Transferase (TdT)
DIG-11-ddGTP	NU-1618-DIGX
Labeling Kits
HighYield T7 Digoxigenin RNA Labeling Kit	RNT-101-DIGX	In vitro Transcription- based random labeling of RNA probes using DIG-11-UTP
HighFidelity Digoxigenin PCR Labeling Kit	APP-101-DIGX	PCR-based random labeling of DNA probes using DIG-11-dUTP
Digoxigenin NT Labeling Kit	PP-310-DIGX	Nick Translation-based random labeling of DNA probes using DIG-11-dUTP
DIG 3'-End Oligonucleotide Labeling Kit	APP-002	3‘-End oligonucleotide labeling with DIG-11-dUTP

Check out complete portfolio of non-radioactive DNA labeling and RNA labeling products as well!

Questions or inquiries?

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Selected References:

[1] McQuaid et al. (1995) A comparison of digoxigenin and biotin labelled DNA and RNA probes for in situ hybridization. Biotech Histochem. 70:147.
[2] Chevalier et al. (1997) Biotin and digoxigenin as labels for light and electron microscopy in situ hybridization probes: where do we stand? J. Histochem. Cytochem. 45:481.
[3] Leary et al. (1983) Rapid and sensitive colorimetric method for visualizing biotin-labeled DNA probes hybridized to DNA or RNA immobilized on nitrocellulose: Bio-blots. Proc. Natl. Acad. Sci. USA 80:4045.
[4] Hoeltke et al. (1990) Non-radioactive labeling of RNA transcripts in vitro with the hapten digoxigenin (DIG); hybridization and ELISA-based detection. Nucleic Acids Res 18 (19):5843.
[5] Morris et al. (1990) Sensitivity of digoxigenin and biotin labelled probes for detection of human papillomavirus by in situ hybridisation. J Clin Pathol 43 (10):800.