1. Cellular penetration and nuclear importation properties of 111In-labeled and 123I- labeled HIV-1 tat peptide immunoconjugates in BT-474 human breast cancer cells 
Bart Cornelissen, Meiduo Hu, Kristin McLarty, Dan Costantini, Raymond M. Reilly 
Nuclear Medicine and Biology 
Volume 34, Issue 1, Pages (January 2007)
DOI: /j.nucmedbio
Copyright © 2007 Elsevier Inc. Terms and Conditions

2. Fig. 1
(A, B) Scheme for the construction of anti-mIgG-tat ICs (anti-mIgG-tat). (i) Anti-mIgG was first reacted with DTPA dianhydride (DTPAA) to introduce DTPA metal chelators. (ii) DTPA–anti-mIgG was then reacted with NaIO4− to oxidize carbohydrates in the Fc domain. (iii) Oxidized DTPA–anti-mIgG was finally reacted with tat peptides, and the resulting Schiff base linkage between tat peptides and DTPA–anti-mIgG was reduced by NaCNBH3. (iv) DTPA–anti-mIgG-tat was labeled with [111In]Cl3 in 200 mM sodium citrate (pH 5.0).
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3. Fig. 2
(A) Size exclusion HPLC chromatogram of [111In]anti-mIgG-tat analyzed on a BioSep SEC-S 2000 column eluted with 100 mM sodium phosphate buffer (pH 6.8) at a flow rate of 1.0 ml/min with detection using a flow-through radioactivity detector. Peaks with retention times (tR) from 5.0 to 6.5 min correspond to [111In]anti-mIgG-tat. The peak with tR=10.0 min is a small amount (<3%) of [111In]DTPA impurity. (B) Relative binding of [123I]anti-mIgG to mIgG coated on a microELISA plate in competition with 6 or 10 nM anti-mIgG or DTPA–anti-mIgG-tat. Values shown are presented as the mean±S.E.M. of replicate measurements (n=5) of binding in the absence of competitors.
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4. Fig. 3
Representative confocal microscopy images of the cellular distribution of DTPA–mIgG (A) or DTPA–anti-mIgG-tat (B). BT-474 cells were incubated with DTPA–mIgG-tat for 2 h at 37°C, fixed, stained with Alexafluor488 anti-goat antibodies (green) and counterstained with DAPI (blue). One-micrometer sections of the cells were imaged. The images shown in (A) were obtained at a magnification of ×630 and show several cells, whereas the images shown in (B) were obtained at a magnification of ×1575 and show a single cell.
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5. Fig. 4
Kinetics of accumulation of radioactivity in BT-474 human BC cells following incubation with 2.7 pmol of [111In]anti-mIgG-tat or [123I]anti-mIgG-tat. (A) Total cellular radioactivity. (B) Cytoplasmic radioactivity. (C) Nuclear radioactivity. Values shown are presented as the mean±S.E.M. of replicate measurements (n=5) of uptake into cells or subcellular compartments expressed as apparent amounts of [111In] or [123I] ICs. The curves were fitted to a one-phase exponential association model by GraphPad Prism Version 4.0 software.
Nuclear Medicine and Biology  , 37-46DOI: ( /j.nucmedbio )
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6. Fig. 5
Accumulation of radioactivity in BT-474 cells following incubation with 2.7 pmol of [111In]anti-mIgG-tat, [111In]anti-mIgG or [111In]DTPA for 1 h at 37°C. (A) Cellular radioactivity. (B) Nuclear radioactivity. Values shown are presented as the mean±S.E.M. of replicate measurements (n=3) of uptake into cells or subcellular compartments expressed as apparent amounts of [111In] ICs or [111In]DTPA.
Nuclear Medicine and Biology  , 37-46DOI: ( /j.nucmedbio )
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7. Fig. 6
Accumulation of radioactivity in BT-474 cells at 1 h at 37°C following incubation with 2.7 pmol of [111In]anti-mIgG-tat in PBS, PBS containing BSA, excess tat peptides or NH4Cl. (A) Cellular radioactivity. (B) Nuclear radioactivity. Values shown are presented as the mean±S.E.M. of replicate measurements (n=3) of uptake into cells or nucleus.
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8. Fig. 7
Kinetics of the efflux of radioactivity from BT-474 cells that imported [111In]anti-mIgG-tat and [123I]anti-mIgG-tat following transfer to a fresh growth medium. (A) Cellular radioactivity. (B) Cytoplasmic radioactivity. (C) Nuclear radioactivity. Values shown are presented as the mean±S.E.M. of replicate measurements (n=5) of retention in cells or subcellular compartments expressed as apparent amounts of [111In] or [123I] ICs. The curves were fitted to a one-phase or a two-phase exponential dissociation model by GraphPad Prism Version 4.0.
Nuclear Medicine and Biology  , 37-46DOI: ( /j.nucmedbio )
Copyright © 2007 Elsevier Inc. Terms and Conditions