98% (Co-migrates with reference on agarose gel electrophoresis)
Concentration:
Minimum 1.6 mg/ml 蛋白
描述:
Human DiI-Labeled Low Density Lipo蛋白
背景:
Purified LDL is labeled with the fluorescent probe, DiI, and reisolated by ultracentrifugation (1.019-1.063). The resultant product is exhaustively dialyzed against phosphate buffered saline, (pH 7.4), sterilized by membrane filtration and then aseptically packaged in a solution containing phosphate-buffered saline at pH 7.4 and 0.2 mM EDTA. Each lot is evaluated on a murine macrophage cell line for fluorescence uptake.
储存和运输:
The labelled LDL is stable for 6 weeks after receipt when handled aseptically and stored at 2-8°C (Don’t Freeze). Note: After prolonged storage, some precipitate may be observed. This is normal for the product. Spin in centrifugation at 5000×g for 10 minutes before using.
Packaging:
The labelled LDL requires one week lead time. Please plan your experiments in advance and use the fresh material.
Native-LDL(n-LDL), Oxidized-LDL (ox-LDL) and -LDL(Ac-LDL) were loaded on agarose gel and electrophoresed for 60 mins. The lipoproteins were stained with Sudan Black (A and B). Oil red O staining was used to determine the formation of foam cell. RAW264.7 were incubated with 80 μg/mL ox-LDL for 24 hrs.
Typical Lipoprotein Labeling Protocol
Dilute DiI-LDL to 10-40 ug/ml in growth media.
Add to cells and incubate for 2-6 hours at 37ºC.
Remove media containing DiI-LDL from your culture.
Wash 3 times with probe-free media.
A. Fluorescence Microscopy:Visualize using standard rhodamine excitation: emission filters (or suggested wavelengths excitation:emission at 554nm:571nm or near). If fixation is desired use 3% formaldehyde in PBS. (Never use methanol or acetone fixation – DiI is soluble in organic solvents). Note: A positive culture must be stained for comparison purposes. A. Cell Sorting:Label as in steps 1-5. Trypsinize or treat cultures with EDTA to produce a single cell suspension. Use labeled pure cultures of positive and negative cell types to set gates on the cell sorter. Suggested Wavelengths for Cell Sorting:Excitation: 554nmEmission: 571nm
