Download File - JOHN J. HADDAD, Ph.D.

Diagnostic Approaches to Maximize Therapeutic Effect 189
sulfide (CdS), cadmium selenide (CdSe), or cadmium telluride (CdTe) and is
insulated by a nonreactive ZnS shell and eliminates photon traps. The mixed
shell of hydrophobic/philic polymer with carboxylic acid derivatization has a
flexible carboxylate surface to which many biological and nonbiological moieties
have been attached. The resulting surface can be attached to antibodies,
streptavidin, lectins, nucleic acids, and related molecules of biological interest to
make them useful for IHC. These novel fluorescent tags have several unique
optical properties that make them very suitable for biological applications that
require multiplexing and highly sensitive molecular detection. The benefits of
quantum immunohistology are that its narrow and symmetric emission reduces
cross talk and facilitates multilabeling. Its high photon output and broad
absorption spectra make possible the use of a single excitation wavelength that
allows for quantitative multiplexing, its high level of brightness and large Stokes
shifts help deal with tissue autofluorescence and finally are 100 more resistant
to photobleaching than organic dyes and makes them ideal probes for fluorescence
quantitation. The slides are stable for long-term archiving. IVD kits are
commercially available. By combining this diagnostic product with appropriate
analytical software, multi-analyte IHC is a reality. Detection and quantitation of
overlapping chromogens accomplished by multispectral imaging is proposed as a
tool that can simplify and enrich the extraction of morphological and molecular
information. Simple-to-use instrumentation is available that mounts on standard
microscopes and can generate spectral image datasets with excellent spatial and
spectral resolution; these can be exploited by sophisticated analysis tools. There
are commercially available, liquid-crystal tunable-filter-based multispectral
imaging platforms. The resulting datasets can be analyzed using spectral
unmixing algorithms to separate out the individual dyes and/or learn-by-example
classification tools. Multiplexed molecular imaging allows the association of
molecular phenotypes with relevant cellular and tissue compartments and conveys
new utility to brightfield-based microscopy approaches.
Genotyping HLA
HLA1 molecules are of major importance for antitumor immune responses.
Expression of HLA1/b2M complexes carrying tumor-specific peptides is a prerequisite
for adaptively matured CTLs to be able to recognize tumor cells (22).
HLA1 are encoded by a family of highly polymorphic genes, with each allele
responsible for a different repertoire of antigen presentation. Thus, even the loss
of a single allele could potentially allow the escape from an antigen-specific
antitumor response. Loss of expression of HLA1 molecules has been frequently
reported in a number of malignant lesions (23). Whether these differences reflect
technical reasons, patient populations’ heterogeneity, and/or the different role
played by HLA1 in various types of malignancies is not known. This would
therefore represent a serious limitation for vaccine-based antitumor therapies. In
the past, the large majority of studies have been performed by immunoperoxidase