Biomedical Research

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Biomedical Research
Reactive Crystallization of Calcium Hydroxyapatite
Alison Mello Department of Chemistry and
Biochemistry University of Massachusetts
Dartmouth
Dapeng Li Department of Materials and
Textiles University of Massachusetts Dartmouth
Chen-Lu Yang Advanced Technology and
Manufacturing Center University of Massachusetts
Dartmouth
Introduction In recent decades, a considerable
amount of research has focused on developing
biomaterials that can both positively react and
replace bone and its derivatives including dentin
and other hard tissues found in teeth. One such
bioceramic is hydroxyapatite (HAP), a calcium
phosphate. Because HAP has been shown to possess
high biocompatibility while causing little or no
damage to existing host tissue, it is currently
used in dentistry and medicine to remodel bone
and as the base or surface coating for a variety
of tooth, bone, and orbital implants. Because
the exact processes of bone formation and
differentiation remain unknown, an understanding
of the properties of its synthetically derived
compliments may be valuable in better mimicking
and manipulating the elusive biochemical
mechanisms in vivo.
(SEM EDX analysis)
Materials and Methods To prepare all aqueous
hydroxyapatite crystals in this procedure, 1.0 M
calcium hydroxide and 0.6 M phosphoric acid were
reacted in a wet crystallization procedure.
Although other calcium and phosphate reactants
will have been shown to produce functional HAP,
the products of the simple acid-base reaction
below are limited to HAP and water, a
biologically non-damaging compound. Crystal
analysis was accomplished using a JEOL JSM 5610
scanning electron microscope (SEM), and energy
depressive X-ray (EDX) analysis was done using an
Oxford 6587 instrument. 10 Ca(OH)2 6 H3PO4 ?
Ca10(PO4)6(OH)2 18 H2O
In all procedures, the calcium hydroxide and
phosphoric acid solutions were reacted in
stoicheiometric volumes so that all of the
calcium hydroxide reacted while little excess
acid was used (Example 50 mL of 1.0 M calcium
hydroxide and 51.0 mL of 0.6 M phosphoric acid).
The excess acid was removed from all products by
washing the filtered samples with at least 300 mL
of distilled water.

• Results to Date
• Fig. 1 shows the EDX microanalysis of calcium
  hydroxyapatite crystals.
• Under SEM analysis, the HAP crystals resulting
  from 60 oC crystallization appear to have a
  slightly more defined crystal structures (Fig.
  2).
• While the crystals of samples at pH 7.1 and 8.5
  appeared similar in shape and size, crystals at
  pH 8.5 appeared to be less scattered throughout
  the surface and have a denser concentration (Fig.
  3).

Fig. 1
(1,000x) at 60 oC
(1,000x) at 27 oC
(500x) at pH 11.5
(1,200x) at pH 8.5
(1,400x) at pH 7.1
Fig. 2
Fig. 3
2
_at_ ATMC

• Results (continued)
• SEM imaging of the samples revealed that crystals
  of the 2.0 bar samples possessed elongated
  oval-like crystals that were uniform in size and
  shape (Fig. 4).
• Fig. 5. shows the consequences of additional
  heating of dry crystal. Qualitative analysis of
  the samples revealed that the additionally heated
  samples were all harder than the control sample.
  Furthermore, the samples heated for 24 hours were
  much harder than their 12 hour, same-temperature
  counterparts, and the samples heated at 150 oC
  were both harder than the crystals heated at 100
  oC. SEM imaging revealed the overall size and
  shape of the crystals in all the over dried
  samples were all similar they all possessed
  surfaces with irregular and non-uniformly shaped
  or sized crystal structures. Novel crystals seen
  in these samples were interspersed within the
  same viewing field. Structures were needlelike,
  fernlike, blossom-shaped, and starburst-patterned.
  

(1,000x) at 1.01 bar
(1,100x) at 2.0 bar
(2,000x) at 2.0 bar
(3,000x) at 2.0 bar
Fig. 4
Fig. 5
(110x) varied crystal structures
(350x) front fernlike pattern
(800x) fernlike crystal
(950x) flower pattern
(2,300x) flower pattern center
(300x) starburst patterns
(300x) starburst pattern
(1000x) flower pattern inverted position
(9,000x) front needlelike crystal
(430x) fernlike, needlelike, and irregular
crystals
(1,000x) needlelike crystals
(700x) needlelike crystals
Dye-sensitized Antibody-nanoparticle Conjugates
for Rapid Detection of Residual Antibiotics
Bin Wu Phosphorex, Inc. Fall River, MA
Chen-Lu Yang Advanced Technology and
Manufacturing Center University of Massachusetts
Dartmouth
Yue Huo Department of Materials and
Textiles University of Massachusetts Dartmouth
Introduction Immunochromatographic strip (ICS)
tests have been applied to more than 100
infectious diseases and more than 60 other
chemical analytes such as hCG, RF, CRP, ASO, FDP,
and fecal occult blood. The popular urine tests
are AMP for amphetamine, BAR for barbiturates,
BZO for benzodiazepines, COC for cocaine, MET for
methamphetamine, MOP for morphine, MTD for
methadone, OPI for Opiate, PCP for phencyclidine,
TCA for tricyclic, and THC for marijuana. Due to
recent reports on misuses and over-uses of
antibiotics by doctors and patients, a need on
urine test for antibiotics is on the horizon.
Excitation
Antibiotics
Nanoparticle
Emission
Antibody
Fluorescent Dye
Fig. 1 The basic principles of dye-sensitized
antibody-nanoparticle conjugation.
3
University of Massachusetts Dartmouth Chapter of
Sigma Xi, the Scientific Research Society 14th
Annual UMass Dartmouth Research Exhibition

• Basic Principles
• By attaching antibody molecules to latex
  nanoparticles, the conjugates are prone to target
  antibiotic molecules through the antibody-antigen
  interaction mechanism (Fig. 1).
• The signal from a single antibiotic molecule can
  be greatly enhanced by hundreds of fluorescent
  dye molecules encapsulated in the
  antibody-nanoparticle conjugate.
• With the enhancement of the dye molecules on the
  conjugates, the detection limit of a fluorometer
  or spectrofluorometer can reach 5 - 50 ppb.
• When properly designed, the latex nanoparticles
  will allow the detection of multiple antibiotics
  on one testing strip.

• Experimental
• Buffer preparation
• Synthesis of color nanoparticles
• Synthesis of fluorescent nanoparticles
• Conjugating nanoparticles to antibiotic
  antibodies
• Confirmation of conjugation

Table 1

• Results
• Table 1 shows the characteristics of color and
  fluorescent nanoparticles.
• Figure 2 shows the nanoparticles of various
  colors.
• Figure 3 shows the Zeta-Plus particle size
  distribution of blue nanoparticles, mean size
  190 nm, polydispersity 0.005.

Fig. 3

• Table 2 shows the particle size of
  antibody-nanoparticle conjugates before and after
  addition of antigen measured by light scattering.
• Figure 4 shows a fluorescent spectrum of the
  fluorescent nanoparticles prepared in this study
  (Entry 4 in Table 1). Rhodamine was the dye used
  in the preparation of the nanoparticles. The
  excitation wavelength was set at 540 nm, the
  emission peak is at 575 nm.
• Figure 5 shows the absorbance of anti-Penicillin
  (200 ml/ml) at 280 nm. It clearly shows the 280
  nm absorption of the antibody.
• Figure 6 shows the UV spectrum of the
  antibody-nanoparticle conjugate suspension in DI
  water.

Fig. 2
Table 2
Anti-Penicillin Control
Anti-P Nanoparticle Conjugates
ABS
ABS
Wavelength (nm)
Wavelength (nm)
Fig. 4
Fig. 6
Fig. 5