Raman Spectroscopy for Early Cancer Detection, Diagnosis and Elucidation of Disease-Specific Biochemical Changes - Emerging Raman Applications and Techniques in Biomedical and Pharmaceutical Fields - page 324

[92], DCIS now forms some 20% of all breast cancers diagnosed by screening.
Approximately 80% of DCIS is picked up due to microcalcifications (see later
in chapter). Once the radiologist has identified suspicious calcifications, these
are biopsied, so that architectural and cytological information can be obtained.
Ductal carcinoma in situ (DCIS) is an intraepithelial proliferation with
features of malignancy localised predominantly within ducts. Studies suggest
up to 50% of patients with microscopic foci of DCIS develop invasive carci-
noma [93]. The invasive lesion occurs in the same area as the original lesion
indicating a likely precursor process [94]. A series of cases in which DCIS
was not excised has been reviewed and these indicate that progression to
invasion is related to the subtype of DCIS: comedo disease progresses into in-
vasive carcinoma both more often and more rapidly than low grade DCIS [95].
Several histopathological systems for subdividing DCIS have been attempted
and there is no clear consensus of opinion between the pathologists [96]. This
causes great concern in the management of these cases [97].

Raman of DCIS

The Raman spectroscopic studies on the breast (outlined above) have demon-
strated discrimination between pathology groups of interest in the breast, but
they have not to date demonstrated a sub-classification of ductal carcinoma
in situ, over which most clinically important decisions are made for early
diagnosis. Here we outline the use of Raman spectroscopic mapping and dis-
crimination for the characterisation and classification of DCIS of the breast.
Breast samples were collected and snap frozen in liquid nitrogen, following
informed written consent during routine breast surgery. The National Coordi-
nation Group for Breast Screening Pathology in the UK recommend a system
derived from the work of Holland and colleagues, which classify DCIS as high,
low, intermediate grade based on cytonuclear features [98]. This study utilised
this system to identify specific ducts of interest for Raman mapping. For each
sample following cryosectioning, an area which corresponded to a marked duct
on the adjacent haematoxylin and eosin (H&E) stained section was selected
for Raman mapping. Figure 13.8 shows an example of an H&E of DCIS and
an image of an unstained duct area for Raman mapping.
A customised Raman microspectrometer [7] was used for this study to en-
able the acquisition of tissue spectra. Laser light at 830 nm was focused on the
tissue section using an Olympus x80 MIRPLAN ultra long working distance
objective to provide 35 mW into a spot of about 3

m in diameter. The
sample was mounted on a Leica DML light microscope, which was equipped
with a motorised, computer-controlled sample stage, which enabled automatic
scanning of the sample. The area to be scanned and the scanning step size
were programmable. Raman maps were acquired using a step size of 10

× 10 μ

m
and a signal collection time for each pixel of 30 s. The selected area varied
from 0 . 4mm 2 (400 spectra) to 4 mm 2 (40,000 spectra) depending on the size
of the ducts imaged.

μ