Fluorescent immunolabeling of cancer cells by quantum dots and antibody scFv fragment

Abstract

Semiconductor quantum dots (QDs) coupled with cancer-specific targeting ligands are new promising agents for fluorescent visualization of cancer cells. Human epidermal growth factor receptor 2/neu (HER2/neu), overexpressed on the surface of many cancer cells, is an important target for cancer diagnostics. Antibody scFv fragments as a targeting agent for direct delivery of fluorophores offer significant advantages over full-size antibodies due to their small size, lower cross-reactivity, and immunogenicity. We have used quantum dots linked to anti-HER2/neu 4D5 scFv antibody to label HER2/neu-overexpressing live cells. Labeling of target cells was shown to have high brightness, photostability, and specificity. The results indicate that construction based on quantum dots and scFv antibody can be successfully used for cancer cell visualization.

Bioconjugation of quantum dots: Review & impact on future application

“…They also designed a scFv-(barnasebarnase) fusion protein (termed 'dibarnase'), in which two barnase units were directly fused head-totail without a linker. This module allowed binding of the scFv Ig to the QDs [61]. Taken together, data has shown that the barnase-barstar system constitutes a useful tool for the design of oligomeric proteins for basic research as well as for imaging [62] and therapeutic applications [60].…”

Feasibility study of the optical imaging of a breast cancer lesion labeled with upconversion nanoparticle biocomplexes

“…6 The cells were cultivated in YTPS broth until the stationary growth phase, and then centrifuged, followed by re-suspension in cold lysis buffer [0.05 M Tris-HCl, 0.1 M NaCl, 10 mM ethylenediaminetetraacetic acid (EDTA), 10 mM dithiothreitol (DTT), pH 8.0]. The obtained solution was sonicated on ice with a 30% ammonium sulfate saturation for cell disruption followed by nucleic acid precipitation with poly (ethyleneimine).…”

“…The UCNP signal in vitro was also modeled using Eqs. (5) and (6), with the optical properties of the phantom and I ex ¼ 100 W∕cm 2 as parameters. As is shown in Fig.…”

“…3 One of the accepted targets for cancer diagnostics is the human epidermal growth factor receptor 2 (HER2/neu), which is overexpressed in a number of cancers and often accompanied by high drug resistance. 6 Identification and precise localization of a cancerous lesion, which is characterized by a high density of HER2/neu using patient-compliant optical means, can aid diagnostic and therapeutic decisions. A recombinant scFv4D5 mini-antibody is an example of such a high-efficiency HER2/neu-targeting vector that represents a single chain variable fragment of immunoglobulin (Ig) and exhibits lower cross-reactivity and immunogenicity in comparison with the corresponding full-size antibody.…”

“…A flexible modular design using a high-affinity molecular linker barstar: barnase (Bs:Bn) was instrumental to achieve bioconjugation of the UCNP to the targeting vector, resulting in a complex assembly of UCNP with scFv4D5 mini-antibody. 6,20 Human breast adenocarcinoma cells SK-BR-3 overexpressing HER2/ neu were target-labeled with the UCNP biocomplexes and imaged using epi-luminescence microscopy. Optical imaging of a compact cancer cell cluster (pertinent to cancer stage I) was simulated using our breast cancer cell model covered with an optical phantom that reproduced live breast tissue properties.…”