Quantifying Polymer Deposition


Table 1: Instrumental parameters used for microfluorometry measurements.


Parameter Green excitation


Objective lens magnification


Photomultiplyer tube voltage1


Diaphragm (measuring element)


Scanning speed Scan length


Description ∼515–560 nm


Lambda max (λ max) ∼600 nm


25× for scans along hair fiber axis; 40× for scans of cross sections


1.0 kV for surface scans; 1.5 kV for scans of cross sections


10 μm× 20 μm for surface scans; 16 μm2


for cross sections


72 μm2 for surface scans; 4 μm2 for cross sections


/sec


∼3 mm for surface scans; 160 μm for scans of cross sections


1For hair fibers the photomultiplier tube voltage was set to 1.0 kV. However, for


cross sections, in order to obtain enough signal the voltage has to be set to 1.5 kV.


an average amount of damage from various environmental factors compared to the root area and the fiber tip. Te fiber ends see the maximum amount of damage since they have full exposure to the environment, whereas the roots of hair fibers see the least amount of damage because the scalp shields them from exposure. Based on this information, hair fibers 2 cm in length were cut from the middle of the hair fibers and were subsequently mounted on glass slides. Prior to treatment with polymer solutions, the tresses


were shampooed using the following protocol: Te tress was wet under running tap water at 1.2 gals/min at a temperature of 37°C. Te hair tress was then rinsed for 30 sec. A quantity of polymer was then weighed out (0.5 ml/gram of hair), applied to the tress, and remained on the hair fibers for 3 min. Te hair tress then underwent a final rinse for 30 sec. Applying the label. Aſter air drying, the samples were


post-labeled with the fluorophore. Te protocol states that the polymer should remain on the hair fibers for approximately 3 minutes since a typical user’s daily grooming practices nor- mally involve leaving conditioner on their hair for no more than several minutes before rinsing. Aſter the hair fibers were treated with the polymer solutions according to the protocol described above, the segments were post-labeled with the fluorophore by immersing them in a 0.025% aqueous solution of sulforhodamine-b solution for 30 sec, followed by rinsing in de-ionized water (DI) water for 10 sec. Te fibers were then air dried and stored in the dark. Preparing cross sections of hair


fibers. Several hair fibers were placed into small plastic molds that were approximately 1 cm in diameter and 2 cm in height. Te procedure involved mounting four or more 1 cm hair fiber segments onto small cardboard sup- ports with a hole cut out near the top.


2019 July • www.microscopy-today.com


Figure 4: (A) Molecular structure of the anionic fluorophore sulforhodamine-b showing the two sulfite (SO3


2–) negative charge sites. Diagram obtained from


Pub Chem open chemistry database. (B) Typical emission spectrum obtained from a hair fiber labeled with sulforhodamine-b. The maximum wavelength was approximately 600 nm.


Individual fibers were laid across the hole vertically, then both ends were glued to the support with five minute epoxy (Figure 5A). Te frame was then inserted into the mold (Figure 5B). Subsequently the molds were filled with a low-viscosity four- component epoxy of medium hardness and placed in an oven to cure overnight at 60°C. Aſter curing, the mold was removed from the oven, and the cast was removed from the mold (Fig- ure 5C). Ten a jeweler’s saw was used to cut out a region where only the embedded hair fibers were visible. Te cast was then mounted on the stage of a Dupont Sor-


vall JB-4 microtome equipped with a diamond knife to create the 10 μm thick cross sections. Te thin sections were subse- quently placed on a glass slide, covered with a drop of immer- sion oil and a glass cover slip.


Figure 5: Photographs showing the cardboard frame with hairs attached (A) that was placed in the plastic capsule (B). Embedded fibers (C, arrows) were visible after the cast was removed from the capsule.


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