恭喜你，打开本文，你将获得一次性解决以上问题的新型WB方法——In-Cell/On-Cell Western

与膜上WB法相比， In-Cell/On-Cell Western更快速便捷

01.Insulin Rearch

Dose-dependent results of human insulin receptor activation. A. Images of ICW signals detected by an Odyssey® Infrared Imaging System (LI-COR, Bad Homburg, Germany) with the 700- and 800-nm channels. The signals obtained from DNA and cell stain with DRAQ5™ and Sapphire700™ excited at ~680 nm and detected at ~700 nm are displayed in red, whereas the signals obtained from detecting the phosphorylated human insulin receptor (p-hIR) excited at ~780 nm and detected at ~800 nm are displayed in green. The composite overlay shows both measurements in one image. 

02.Signaling Pathways

Effects of Y27632 and fasudil on p-ERK protein expression levels in cultured spinal cord microglia.(A) In-cell western blot assay of the changes in p-ERK protein expression levels in response to Y27632 (10 μM) and fasudil (41 μM) treatments at 1, 3 and 6 hours. (B) Statistical analysis of p-ERK protein levels. [1]

03.Drug Rearch

Tim-3 is mostly located inside resting THP-1 cells but is externalised after PMA-dependent PKCα activation. Tim-3 was measured using a Li-Cor on cell assay with (in cell western) or without (on cell assay) methanol permeabilisation (see the Materials and methods section). Exposure to the anti-Tim-3 antibody was performed for 2 h. Images are from one experiment representative of five which gave similar results.[2]

04.siRNA

Representative in-cell western dual detection of K18 protein (green), β-actin protein (red), and combined (merged image-yellow) following exposure to either Lipofectamine™ or equimolar concentrations of a non-targeting siRNA (siCTL) as controls, or 100 nM siRNA to KRT8/18 (siKRT8/18). Diminished expression of K18 in the single and merged images was evident following 72 h exposure to siKRT8/18.[3]

05.Apoptosis

Following the pre-incubation of HK-2 cells with 10, 20 and 40 µM SchB for 2 h, the cells were stimulated with cis-DDP for 24 h. The activation of caspase-3 was then determined by in-cell western blot analysis. [4]

06.Ion Channel

Detection of P‐Akt, the active (phosphorylated) form of Akt within TH+ neurons using infrared fluorescence imaging applied to the In‐Cell western blot technique in 7 DIV cultures exposed chronically to GLIB in the presence or not of LY applied only acutely (20 min). [5]

In-Cell/On-Cell Western 应用广泛

参考文献

1. The Rho-associated kinase inhibitors Y27632 and fasudil promote microglial migration in the spinal cord via the ERK signaling pathway[J]. Neural Regeneration Research, 2018, v.13(04):111-117.

2. Yasinska I M , Ceccone G , Ojea-Jimenez I , et al. Highly specific targeting of human acute myeloid leukaemia cells using pharmacologically active nanoconjugates[J]. Nanoscale, 2018, 10.

3. Trisdale S K , Schwab N M , Hou X , et al. Molecular manipulation of keratin 8/18 intermediate filaments: modulators of FAS-mediated death signaling in human ovarian granulosa tumor cells[J]. Journal of Ovarian Research, 2016, 9(1):8.

4. Liu Q , Song J , Li H , et al. Schizandrin B inhibits the cis DDP nduced apoptosis of HK2 cells by activating ERK/NF?κB signaling to regulate the expression of survivin[J]. International Journal of Molecular Medicine, 2018.

5. Toulorge D , Guerreiro S , Hirsch E C , et al. KATP channel blockade protects midbrain dopamine neurons by repressing a glia-to-neuron signaling cascade that ultimately disrupts mitochondrial calcium homeostasis[J]. Journal of Neurochemistry, 2010, 114(2):553-564.

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