Y. Yang et al. / Carbohydrate Research 361 (2012) 189–194
193
values calculated from Langmuir 1:1 fitting model were not reli-
able. Although SPR has been used in investigating carbohydrate–
protein interactions, the glycan is rarely placed in the flow buffer
because of the low sensitivity induced by the low molecular weight
of the sugar. On the other hand, it is very difficult to accurately
measure fast kinetics30 which appear in the interaction between
free sugar and PNA. To overcome this problem, Yamanoia and co-
ZnS QDs displayed a PL emission (kem = 534 nm) with the full
width at the halfmaximum (FWHM) less than 30 nm.
Orange QDs (kem = 564 nm) were synthesized with the same
procedure except for changing the ratio of the CdO (0.26 g), oleic
acid (2.5 ml) Se (12 mg), and S (192 mg). The product is a red
power.
Red QDs (kem = 591 nm) CdO (0.26 g), oleic acid (2.5 ml), Se
(16 mg), and S (192 mg). The product is a black power.
workers.31 synthesized
D-galactose-b-cyclodextrin conjugates and
placed the sugar in the flow phase, the KD value they got was
1.08 ꢀ 10ꢁ5. Jayaraman et al.32 synthesized aminoethyl lactose
and immobilized the sugar on CM5 chip, they also observed the
binding and calculated the KD value (1.46 ꢀ 10ꢁ4).
4.2. Preparation of GalSH, LacSH, and GlcSH
GalSH, GlcSH, and LacSH were synthesized according to the pro-
cedure reported before.18,19
Langmuir 1:1 fitting model was used to calculate the KD value in
the multivalent system. It is important to note that although we
determined the formulas of the glyco-QDs from ICP-OES and
NMR results, they were only estimations. Therefore, the molar con-
centration could not be accurately calculated. Moreover, the algo-
rithms for SPR analysis were developed only for bivalent systems29
and the precise molar ratio of ligand to target is not known. As a
result, the binding parameters like kd and ka cannot be accurately
measured.
4.3. Surface plasmon resonance Analysis
The SPR results were obtained on the BIAcoreTM 3000 instru-
ment (BIAcore, Uppsala, Sweden). Lectin-functionalized BIAcore
sensor chips were prepared from carboxymethylated sensor chips
(CM5, BIAcore) by NHS/EDC activation followed by injection of PNA
in acetate buffer (10 mM, pH 4.5).
An enhancement factor b (Kfree/Kmultivalent)
33 for error reduction
is presented to define the increased affinity of a multivalent inter-
action. The b of the free sugar was defined as 1 (Table 2). For the
Gal-QDs and Lac-QDs, multivalent attachment of the weakly bind-
ing sugar enhanced avidity toward the protein. The b values indi-
cated the interactions between protein and Gal-QDs and Lac-QDs
are 3.1 ꢀ 106 and 1.5 ꢀ 107 fold higher than that of the free sugar,
respectively. PNA was sensitive to increased sugar valency and a
relative potency per galactose of 2.3 ꢀ 104 and lactose of
1.4 ꢀ 105 was reached.
Acknowledgments
This work was financially supported by the Natural Science
Foundation of China (Grant No. 90713004, 20732001), the State
New
Drug
Innovation
(Grant
Nos.
2009ZX09103-044,
2009ZX09501-011), the National Basic Research Program of China
(Grant Nos. 2012CB822100), and the State Key Laboratory of Nat-
ural and Biomimetic Drugs and Peking University.
Supplementary data
3. Conclusion
Supplementary data associated with this article can be found,
We synthesized QDs with different emission wavelengths and
coated them with different sugars. The formulas of the glyco-QDs
were determined by NMR and ICP-OES. SPR analysis showed that
the lectin specificity was maintained and the affinity was enhanced
after coating on QDs. These multicolor fluorescent probes can be
used to label glycoproteins and cells in the study of carbohy-
drate–protein interactions and glycoprotein functions.
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CdO (0.26 g), oleic acid (OA, 2.5 ml), and tri-n-octylamine (TOA,
15 mL) were mixed in a three-necked flask and heated to 300 °C
under an argon atmosphere to yield a clear solution. A stock solu-
tion of Se (6.4 mg) and S (0.192 g) in 3.0 mL TOP was swiftly in-
jected into the hot solution, and the reaction was allowed to
proceed at 280 °C for 1 min. ZnS stock solution (a mixture of
0.2 mM ZnO powder in 1.0 mL oleic acid and 0.2 mM S powder in
1.0 mL TOP) was then added dropwise (1 drop per second) at
220–240 °C. CdSeS/ZnS-QDs were obtained with ethanol sedimen-
tation and repeatedly washed with ethanol. The prepared CdSeS/