Figure 4A shows a different set of biopsy samples visualized under white light following treatment with the AF350-WGA probe. The fluorescent lamp used for white selleck products light imaging may have caused uneven tissue illumination, resulting
in the cancerous tissue looking brighter in Figure 4A. However, tissue appearance differences between normal and diseased tissue is well established due to increased cell density, protein amounts, etc. Typically, these lesions are often times whiter in appearance which would have caused them to appear brighter under white light imaging. Nevertheless, increased probe fluorescence is noted on the tumor specimen and not the normal specimen ( Figure 4B), proving the specificity of the probe for the overexpressed glycan residues on the tumor surface. Lastly, Figure 4C shows a digital camera image of tissue biopsies incubated in AF350-WGA to capture fluorescent images that would more accurately demonstrate the conditions observed within a clinical setting; this image shows the enhanced fluorescence is easily visible
with the naked eye. Similar results were seen for all tissue samples tested with AF350-WGA and are summarized in Figure 5 and in Table 2. Figure 5 shows the patient/tissue samples’ SNR for AF350-WGA testing. The AF350-WGA fluorescence of the cancerous tissue was statistically significantly higher than that of normal tissue with an average SNR of 5.88 ± 3.46 (P Nutlin-3a molecular weight = .00046, Table 2). The differences observed amongst the SNRs can be attributed to the fact that sialic acid overexpression is dependent on patient variability, disease progression, cancer aggressiveness, etc. However, it is important to note that all patients displayed SNRs greater than 3. The UV autofluorescence of the cancerous tissue displayed an average SNR of 1.35 ± 0.41 and was not statistically significantly triclocarban different than normal tissue (P = .098, Table 2). The SNR of AF350-WGA was statistically significantly larger than the SNR for UV autofluorescence (P = .0049, Table 2) with it being at
least double the ratio in all seven patients. To further validate the specificity of the WGA binding conjugate, inhibitory experiments were carried out with N-acetyl glucosamine which serves to block the available binding sites of WGA prior to sample application. Pre-incubation of AF350-WGA with the sugar resulted in a threefold decrease in fluorescence intensities of the cancerous tissue (Figure 6), indicating that the soluble sugar competitively inhibited the WGA from binding to the overexpressed glycan residues on the cancerous cell surface. Interestingly, the inhibited AF350-WGA still resulted in higher fluorescence intensity values from the cancerous tissue when compared to the normal tissue (Figure 6B and C).