The Standard in Quantitative Western Blots
Figure 1. Serial dilutions (10 ng to < 1 pg) of purified human transferrin (Tf) were used to assess Western sensitivity. The Odyssey Classic using infrared fluorescence detection, reproducibly detected 1.2 pg of Tf, while only 4.9 - 9.5 pg was detected with chemiluminescence. Infrared detection sensitivity was approximately 200-fold greater than previous studies with visible fluorophores (Cy®3, Cy®5, or FITC).
Go Green with Infrared Imaging - Cleaner Planet, Better Data
No Darkroom, No Harsh Chemicals
Infrared imaging on the any of the Odyssey® imaging systems eliminates the need for film and the harsh chemicals and excessive water use associated with film development.
In addition to the environmental advantages of infrared imaging, it has also become the industry leader in Western blot technology since LI-COR pioneered the field more than 10 years ago. When used with corresponding infrared fluorescent-labeled secondary antibodies, LI-COR infrared technology offers the following benefits:
Quantitative analysis and a wide linear dynamic range that is not available with traditional chemiluminescent methods
Detect strong and weak bands on the same blot, without "blown out" lanes or hidden bands (Fig. 1)
Detect two targets simultaneously on the same membrane to increase quantification accuracy
Infrared dyes offer advanced signal stability that allows for convenient and reproducible data that are not time-sensitive - data are not contingent on the lifespan of an enzymatic reaction (Fig. 2)
Figure 2. Western blots containing purified transferrin were processed using chemiluminescence (10 ng - 1.2 pg) or near-infrared detection (40 ng - 4.8 ng). Panel A shows the loss of signal for chemiluminescence detection after 60 minutes on the Odyssey Fc. Panel B shows the stability of a Western blot processed with IRDye 800CW secondary antibody after 18 months imaged on the Odyssey Classic. The lower limit of detection is maintained in the near-infrared Western blot with all of the data points available (14 points total) even after storage for 18 months. Compare this to the chemiluminescence detection where 7 of the same data points are detected after only 60 minutes.
Consider the following characteristics associated with film development and its required reagents:
Photo solutions have high chemical and biological oxygen demand (COD and BOD) and are often treated with ozone, peroxide, or aeration to reduce COD.
Photo chemicals use non-biodegradable compounds in developers and washing aid solutions, such as EDTA, DTPA, NTA, and borate. Water containing these chelating agents can leach metal from water treatment equipment, as well as pipes.
Many darkrooms use potassium ferricyanide, potassium dichromate, or permanganate for bleaching. These compounds decompose in the wastewater streams and are highly regulated for sewer disposal, at a dilution of 20,000 times or more.
Hydroxylated benzene compounds are commonly used as developing agents. They are harmful to humans and experimental small animals, and have been noted as mutagenic. These compounds have a high COD.
