Page 100 - Vibrational Spectroscopic Imaging for Biomedical Applications

76 Cha pte r T h ree

(a) Adipocytes in Growth Medium (d) 2 3 (i) TAG Reference
υ (CH ) 8
w
2
9
υ w (CH ) υ s (CH ) 10
3 3
υ s (CH ) 7
2 6
11
Lipid Micelles
3100 3000 2900 2800 1800 1600 1400 1200 1000 800
2 (ii) Formalin - Water Rinse - Air-Dried
(b) Formalin - Water Rinse - Air-Dried
Lipid Deposit
3
9
8 10
1 4 6 7
Lipid Deposit 3100 3000 2900 2800 1800 1600 1400 1200 1000 800
2
(c) PF - OsO - CPD Fixed (iii) PF - OsO 4 - CPD Lipid Micelles
4
3
Lipid Micelles 4 5 6 89 10 11
7
3100 3000 2900 2800 1800 1600 1400 1200 1000 800
–1
Wavenumber (cm )
(e) CH O
H CH 3 3 O
C C Os O
Scheme I + OsO 4 H
C C O
H CH 3 H
CH 3
H C H C H C CH 3
3
3
3 O O
CH CH CH CH
Scheme II + 2OsO + Os O + OsO
HC 4 HC 3
CH CH
O O
CH 3 CH 3 H C CH 3
3
FIGURE 3.7 Optical photomicrographs showing (a) adipocytes in growth
medium with prominent intracellular droplets (scale bar = 10 μm);
(b) adipocyte following formalin fi xation, water rinsing and air-drying (scale
bar = 20 μm); (c) adipocytes fi xed in paraformaldehyde (PF) and osmium
tetroxide (OsO ) and critical-point-dried (CPD) (scale bar = 30 μm); (d) typical
4
FTIR spectrum of (i) triacylglyceride (TAG) reference with C to C saturated
2 10
hydrocarbon chains, (ii) the lipid deposit from a formalin-fi xed air-dried
adipocyte and (iii) the lipid droplets from a PF and osmium tetroxide (OsO )
4
CPD adipocyte; (e) OsO reaction with unsaturated hydrocarbon chains to
4
form cyclic esters. Scheme I, reaction with a single unsaturated hydrocarbon
chain. Scheme II, cross-linking reaction with adjacent unsaturated
hydrocarbon chains.
peaks labeled 8 to 10. However, the peaks in this spectrum [Fig. 3.7d(ii)]
are broader compared with the same peaks in the reference TAG spec-
trum [Fig. 3.7d(i)]. This is due to collapse of the lipid droplets that give
rise to a range of bonding strengths with neighboring molecular spe-
−1
cies for those functional groups absorbing at frequencies >1500 cm .