TLectinSense on the ASK

• Overview
• Products and catalog numbers
• Using TLectinSense in vivo/ex vivo
• Using TLectinSense in vitro
• Application notes and posters
• Citations

Overview

The protein tomato (L. esculentum) lectin is widely used for vascular labeling because the protein has high binding affinity for glycoprotein N-acetylglucosamines on the surface of vascular endothelial cells. TLectinSense™ 680 is a near-infrared fluorescent imaging probe which is comprised of the tomato lectin protein and a near-infrared fluorophore. TLectinSense 680 targets the vasculature and enables imaging of blood vessels and angiogenesis. It was designed and optimized for use in living animals. It can be used in assessing vascularity in vivo and in real time, without termination of mice, excision and processing of the tissue. It can also be used for improving the efficacy, early detection and monitoring of anti-angiogenic therapies.

Tumors induce significant blood vessel development in order to support their aggressive growth and progression. The extensive and disordered nature of tumor vasculature impairs drug delivery and efficacy. Therefore, destroying the tumor vasculature and/or inhibiting neo-vascularization have become well-accepted and proven cancer treatment strategies. TLectinSense 680 targets the surface of vascular endothelial cells and can be used for quantitation of vascular burden across different tumor cell lines.

Figure 1: TLectinSense 680 mechanism of action. A near infrared fluorophore (VivoTag 680 XL) was conjugated to the tomato lectin protein to produce the imaging agent.

Figure 2: Tomato lectin is a recognized approach to labeling the vascular endothelium, intestinal epithelium and microglia. The mechanism for targeting vascular endothelium with tomato lectin is an established gold standard.

Products and catalog numbers

Using TLectinSense in vivo/ex vivo

The recommended procedure for in vivo imaging with TLectinSense 680 is intravenous administration and imaging 6 hours post injection.

Imaging in matrigel plugs and tumor models: TLectinSense 680 can be used to study angiogenesis and blood vessel density, in matrigel plugs and animal tumor models.

View instructions on setting up an in vivo mouse experiment with TLectinSense 680 and imaging on an IVIS or FMT system.

Figure 3: A) LLC tumor-bearing mice were injected with TLectinSense 680, imaged tomographically at 6 hours and fluorescence was quantified in 5 different organs. Tissues were then collected and fluorescence assessed by planar imaging (ex vivo). Mean counts/energy for each tissue were determined as a measure of tissue brightness. B) The plasma pharmacokinetic profile was assessed by injecting CD1 mice (3/time point), collecting plasma at multiple times post-injection and measuring plasma fluorescence.

In Vivo/Ex Vivo Imaging

Figure 4: A) Lewis Lung Carcinoma cells were injected into the right flanks of Nu/Nu female mice and grown for 10 days. Mice were imaged tomographically (FMT 2500LX) 6 hours after intravenous TLectinSense 680 injection (2 nmoles; three representative mice are shown). B) Quantification of the tumor and contralateral site signal (pmoles) and C) Shows localization of the fluorescent signal in a frozen tumor section (Blue: DAPI nuclear staining, Red: TLectinSense 680, Green: anti-CD31).

Ex Vivo imaging

Figure 5: Labels vasculature in normal tissues. In vivo administration of TLectinSense 680 also shows labeling of the liver and muscle vasculature, as confirmed by ex vivo co-localization with Anti-CD31-FITC.

Using TLectinSense in vitro

TLectinSense 680 enables imaging of endothelial glycoproteins in endothelial cells such as HUVEC.

Flow cytometry and in vitro microscopy

We have validated TLectinSense 680 for use with fluorescence microscopes and flow cytometers. Here is a brief protocol with a recommended concentration of agent to use:

1. Culture cells in standard TC plate or chamber slide. 
2. Incubate cells with 0.25 uM TLectinSense 680 for 30 min at 37 °C.  
3. Wash 2x with PBS. For flow cytometry, detach and resuspend cells in PBS. 
4. Flow cytometry filter settings: 712/21
   Fluorescence microscopy filter: Cy5.5

In Vitro Imaging:

Figure 6: Human umbilical vein endothelial cells (HUVEC) were incubated with TLectinSense 680 in the absence (TL680) or presence (Competition) of 50X unlabeled lectin and visualized by fluorescence microscopy. DAPI nuclear staining is in blue and TLectinSense 680 signal is shown in red.

Figure 7: TLectinSense 680 binds to human umbilical vein endothelial cells (HUVEC). The effect of plasma on binding was tested by incubating the agents (1 mM or 20 mg/ml) in either PBS or 100% FBS. As a control, commercially-available Lectin-FITC (20 mg/ml) was used. In blue: DAPI nuclear stain, red: TLectinSense 680, Green: Lectin-FITC. Binding of TLectinSense 680 to HUVEC is not inhibited by plasma (FBS), whereas binding of commercial lectin-FITC is inhibited.

Application notes and posters

• Poster: Targeted in vivo imaging of tumor vasculature using a near infra-red labeled tomato lectin agent

Citations

Please visit our Citations Library for references using TLectinSense 680 on the IVIS or on the FMT.