Highly Sensitive Single Domain Antibody–Quantum Dot Conjugates for Detection of HER2 Biomarker in Lung and Breast Cancer Cells

Despite the widespread availability of immunohistochemical and other methodologies for screening and early detection of lung and breast cancer biomarkers, diagnosis of the early stage of cancers can be difficult and prone to error. The identification and validation of early biomarkers specific to lung and breast cancers, which would permit the development of more sensitive methods for detection of early disease onset, is urgently needed. In this paper, ultra-small and bright nanoprobes based on quantum dots (QDs) conjugated to single domain anti-HER2 (human epidermal growth factor receptor 2) antibodies (sdAbs) were applied for immunolabeling of breast and lung cancer cell lines, and their performance was compared to that of anti-HER2 monoclonal antibodies conjugated to conventional organic dyes Alexa Fluor 488 and Alexa Fluor 568. The sdAbs–QD conjugates achieved superior staining in a panel of lung cancer cell lines with differential HER2 expression. This shows their outstanding potential for the development of more sensitive assays for early detection of cancer biomarkers

Assessment of the volume of SWCNT aggregates in the lung tissue sections from w/t and MPO k/o mice.

<p>a. Representative images of the lung tissue sections. Insert - higher magnifigation (2.5× zoom) of a field illustrating the presence of SWCNT (green punctuate spots pointed by white arrows). b. Quantitation of SWCNT aggregates (SWCNT volume/total lung volume) using their specific absorbance (750–850 nm), * p<0.05, <i>vs</i> w/t 1 day post exposure, # p<0.05, <i>vs</i> w/t 28 days post exposure. c. Assessment of SWCNT aggregates - number per microscopic field - using an automated IN Cell Analyser 1000 microscope, * p<0.05, <i>vs</i> w/t 1 day post exposure, # p<0.05, <i>vs</i> w/t 28 days post exposure.</p

Characterization of pulmonary inflammatory responses to SWCNT in w/t and MPO k/o mice at day 1 after pharyngeal aspiration exposure.

<p>a–c. Levels of pro-inflammatory cytokines (a - TNF-α; b - IL-6; c – MCP-1) in BAL fluid of w/t and MPO k/o mice. Mean ± SEM (n = 6 mice/group). *p<0.05, <i>vs</i> control PBS-exposed mice. d. Content of PMNs in BAL fluid of w/t and MPO k/o mice. Mean ± SEM (n = 6 mice/group). *p<0.05, <i>vs</i> control PBS-exposed mice. e. Typical microscopic images of inflammatory cells in BAL fluid with SWCNT inclusions (red arrows). f. Content of PMNs with engulfed SWCNT in BAL fluid of w/t and MPO k/o mice. Mean ± SEM (n = 6 mice/group). *p<0.05, <i>vs</i> w/t mice.</p

Changes in the content of collagen and average thickness of alveolar connective tissue in the lungs of w/t and MPO k/o mice at days 1 and 28 after pharyngeal aspiration of SWCNT.

<p>a. Accumulation of collagen in the lung of w/t or MPO k/o mice. Mean ± SEM (n = 6 mice/group). *p<0.05, <i>vs</i> control PBS-exposed mice, # p<0.05, <i>vs</i> w/t mice 28 days post exposure. b. Morphometric assessments of average thickness of alveolar connective tissue in the lung of w/t or MPO k/o mice. Mean ± SEM (n = 6 mice/group). *p<0.05, <i>vs</i> control PBS-exposed mice, # p<0.05, <i>vs</i> w/t 28 days post exposure.</p

Evaluation of SWCNT size distribution in solubilized lungs of w/t and MPO k/o mice at days 1 and 28 post exposure by t

<p>ransmission electron <b>microscopy.</b> a. Typical TEM images of SWCNT after solubilization of the lung tissue. b. Size distribution of SWCNT present in the lung at days 1 and 28 post exposure. * p<0.05, <i>vs</i> w/t 1 day post exposure, # p<0.05, <i>vs</i> w/t 28 days post exposure. c. Changes in size distribution of SWCNT at day 28 post exposure compared to day 1 post exposure.</p