ROTI®Lumin, 2 x 120 ml
REF P078.1
€ 539,50
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ROTI®Lumin, 2 x 120 ml
Información del producto
P-frases: P210 P280 P302+P352 P304+P340 P305+P351+P338 P308+P311
H-frases: H225-H290-H302+H312+H332-H318-H336-H370
Nivel de riesgo: 3 (8) II
Código hs: 38229000
Marca: Carl Roth
ROTI®Lumin, 10x conc., fluorecent-free, for protein and cytoimmunochemistry, Storage temp. +4 °C
- Sensitive and reliable
- For all standard membrane types
- Low background
- No cumbersome preparation of peroxidase buffers
- Suitable for multiple stripping and hybridisation
ROTI®Lumin is a kit for chemoluminescent detection of peroxidase-linked reporters (e.g. HRP-antibodies), as can be found in Western-Blot assays. In comparison to chromogen substrates, ROTI®Lumin displays increased sensitivity and reduced background reaction.
Mechanisme
ROTI®Lumin is transformed by the HRP into an excited dianion in the presence of hydrogen peroxide. When the dianion is reversed back to its normal state, light is emitted. The light emission of ROTI®Lumin reaches its maximum within 5 minutes and remains at the maximum level for approx. 1-2 hours.
ROTI®Lumin 10x conc., fluorescence-free, for protein and cytoimmunochemistry
Gebruiksinstructies
ROTI®Lumin consists of two solutions, which should be mixed at a 1:1 ratio before use. It can be used with nitrocellulose, nylon and PVDF membranes. Optimal results can be achieved with Immobilon-P® PVDF or ROTI®PVDF.
Can be used for Western-Blotting, ELISA, Southern- Blotting, Dot-Blotting and colony hybridisation. Membranes may be stripped from bound probes using ROTI®Free stripping buffer, and may thus undergo multiple reprobing.
Figure: Sensitivity of the ROTI®Lumine.
Sensitivity of ROTI®Lumin ultra is found in low femtogram range. Detection of a polyclonal antibody (left to right 4 pg, 400 fg, 40 fg, 4 fg) on ROTI®PVDF (Art. No. T830.1). Staining in PBST via a HRP-conjugated anti-rabbit antibody (1:2000).
a/A: ROTI®Lumin ultra; b/B: ROTI®Lumin plus; c/C: ROTI®Lumin; d/D: Product of a competitor, recommended for femtogram range. a/b/c/d: 10 sec. exposition, A/B/C/D: 1 min. exposition.
Sensitivity of ROTI®Lumin ultra is found in low femtogram range. Detection of a polyclonal antibody (left to right 4 pg, 400 fg, 40 fg, 4 fg) on ROTI®PVDF (Art. No. T830.1). Staining in PBST via a HRP-conjugated anti-rabbit antibody (1:2000).
a/A: ROTI®Lumin ultra; b/B: ROTI®Lumin plus; c/C: ROTI®Lumin; d/D: Product of a competitor, recommended for femtogram range. a/b/c/d: 10 sec. exposition, A/B/C/D: 1 min. exposition.
Figure:
Typical course of light emission of the ROTI®Lumins: 0-30 mins., post mixing/spraying. Competitor P: Product of a competitor, recommended for femtogram range.
Typical course of light emission of the ROTI®Lumins: 0-30 mins., post mixing/spraying. Competitor P: Product of a competitor, recommended for femtogram range.
De kit bevat
120 ml (25 ml) ROTI®Lumin 1 (Art. No. P079) and 120 ml (25 ml) ROTI®Lumin 2 (Art. No. P080).
Contents of this Kit may not be bought separately.
Figure:
Relative sensitivity of ROTI®Lumins. Measurement of the relative light emission following ECL reaction with solubilised HRP.
Competitor P: Product of a competitor, recommended for femtogram range.
Relative sensitivity of ROTI®Lumins. Measurement of the relative light emission following ECL reaction with solubilised HRP.
Competitor P: Product of a competitor, recommended for femtogram range.
Figure:
Typical course of light emission of the ROTI®Lumins: 0-5 mins., post mixing/spraying. Competitor P: Product of a competitor, recommended for femtogram range.
Typical course of light emission of the ROTI®Lumins: 0-5 mins., post mixing/spraying. Competitor P: Product of a competitor, recommended for femtogram range.
Technische informatie
| Toepassing | Western, in situ, Histochemistry |
| Enzyme | Horseradish Peroxidase |
| Bewijs | chemoluminescent |
| Signaalproduct | Precipitate (Blot) |
| Toepassing | Western |
Assay protocol for use of BCIP/NBT in immunoblot procedures:
Stock solutions (All solutions are stable for at least 1 year at 4 °C):
0.5 g NBT in 10 ml 70 % dimethylformamide;
0.5 g BCIP (p-toluidine salt) in 10 ml 100 % dimethylformamide.
Incubation buffer for alkaline phosphatase:
100 mM NaCl, 5 mM MgCl2,100 mM Tris (pH 9.5).
Fresh substrate solution: 66 μl NBT stock solution + 10 ml incubation buffer, mix well, add 33 μl BCIP stock solution. Use within 1 hour.
Blot development: Approx. 10 ml substrate solution per 15 x 15 cm2 membrane surface. Develop at room temperature until bands become visible (approx. 30 mins).
Reaction stop: Rinse with PBS/20 mM EDTA.
Detection of peroxidase activity (Immunoassay):
Dissolve 1 mg TMB in 0.1 ml dimethylsulfoxide (Art. No. 4720); add 9.9 ml of a 0.1 M sodium acetate solution (pH 6.0) (Powder: Art. No. 6779), filter and add H2O2 (Art. No. 8070) (final concentration 0.01 %).
Always prepare freshly!
Incubation 10-30 mins at room temperature (approx. 50 μl per microtitre well; finally add 50 μl 1 M H2SO4, Art. No. X873, per well).
Photometric quantitation at 450 nm.
Reference: Bos E.S. et al. (1981) J. Immunoassay 2, (3/4), 187.
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