2.7 Ice crystal size analysis: bright-field microscopy
Ice cream containers were transferred from the −25 °C walk-in freezer to a −18 °C
storage room 1 h before microscopy analysis. All mechanical devices required for
further treatment of the samples were precooled to −18 °C. A thin slice of ice cream
was cut from the core section at the center of the container using a sharp knife. This layer
was placed on a drop of iso-amyl-butanol (previously cooled to −18 °C) on a standard
glass microscope slide, and a few more drops of iso-amyl-butanol were added to the
sample. The microscope slide was covered with a coverslip and placed above liquid
nitrogen in an insulated Styrofoam container and immediately transported to the cold
stage (Linkam Scientific Instruments, Surrey, UK), which was previously programmed
to a constant temperature of −17 °C (Flores and Goff 1999a).Images from uncycled and cycled samples were acquired using an Olympus BH
PLM light microscope. Several different fields were photographed from 2 different
containers of ice cream to obtain at least 300 crystals per sample. These images were
analyzed using the Image J Software (Version 1.46k, Wayne Rasband, National
Institutes of Health, Bethesda, MD, USA) by manually tracing the perimeter of ice
crystals with a computer mouse; the area of each crystal was automatically calculated
by the software. Equivalent circular diameter of the crystals was determined using
Microsoft Excel 2007. Logistic dose–response model with a cumulative distribution
of equivalent circular diameters was used to distinguish ice crystal size distributions.
The slope of the distribution (n) and the X50 values were calculated by
Curve Expert 1.3 program. The X50 value corresponds to the theoretical median
value of the fitted data to the model at 50% of the cumulative distribution. The
276 M. BahramParvar, H.D. Goff
slope parameter is an indication of the span of the population with lower slope
representing a more narrow distribution.