Biomedical subjects

M Rusckowski

Publications and source records attributed to M Rusckowski.

At least 37 records · Page 2Linked to original sources

PubMed1997

Preparation and use of NHS-MAG3 for technetium-99m labeling of DNA.

The chelator mercaptoacetylglycylglycylglycine (MAG3) is on of several amidothiols that have been used successfully to radiolabeled proteins and other molecules with 99mTc. Prior to radiolabeling, the sulfur in these amidothiols is usually protected by a benzoyl group (i.e. S-benzoyl MAG3) which requires extreme alkaline pH or boiling water temperatures for rapid deprotection. As a result, the benzoyl-protected chelator is radiolabeled prior to conjugation (i.e. preconjugation labeling) in the case of carriers such as proteins or polypeptides which cannot withstand harsh conditions. We have employed a simple, two-step, synthesis of the N-hydroxysuccinimide ester of MAG3 in which the sulfur is protected with an acetyl group (i.e. S-acetyl NHS-MAG3). A single-stranded amine-derivitized DNA was coupled with NHS-S-acetyl MAG3. Radiolabeling was accomplished at room temperature and neutral pH by transchelation from 99mTc-tartrate. In comparison to labeled SHNH-DNA, the labeled MAG3-DNA was unstable to cysteine transchelation, however, in contrast to SHNH-DNA, no evidence for serum protein binding of the labeled MAG3-DNA was observed. We conclude that the S-acetyl NHS MAG3 bifunctional chelator may prove to be an attractive alternative method of radiolabeling DNA and other biologically important molecules with 99mTc.

Chelating Agents↗

PubMed1997

Effect of endogenous biotin on the applications of streptavidin and biotin in mice.

The use of streptavidin-conjugated antibody to pretarget tumors in animals and patients, prior to administration of radiolabeled biotin, has provided encouraging results, in part because of the high affinity of biotin for streptavidin and the rapid whole-body clearance of biotin. However, binding of endogenous biotin to streptavidin may interfere with the clinical potential of this approach. This report evaluates the effect of endogenous biotin on an antibody-streptavidin conjugate in a mouse tumor model. Tumored nude mice were depleted of endogenous biotin by sequential intraperitoneal injections of streptavidin. The assay of serum biotin levels indicated less than 0.5 ng of biotin per mL of serum in treated mice versus 4 ng per mL in untreated animals. Flow cytometric analysis was used on single-cell suspensions of tumor from animals receiving streptavidin-conjugated IgG to detect the presence of the antibody on the cell membrane (with fluoroisothiocyanate-conjugated goat anti-mouse antibody), and to detect biotin binding sites on streptavidin (with biotin-phycoerythrin). Both treated and untreated mice demonstrated the presence of antibody on tumor cells through 48 h postadministration, but only in treated animals were biotin binding sites observed. These results in the mouse model suggest that the small concentration of streptavidin delivered to a tumor via a specific antibody may be saturated with endogenous biotin and therefore not able to be targeted subsequently with radiolabeled biotin.

Animals↗

PubMed1997

Technetium-99m labeled epidermal growth factor-tumor imaging in mice.

We have shown previously that the epidermal growth factor peptide (EGF) may be radiolabeled with 99mTc at room temperature and neutral pH by using the N-hydroxysuccinimide ester of S-acetyl mercaptoacetyltriglycine (MAG3) as a bifunctional chelator. By a competition binding assay, we found that MAG3-conjugated EGF retained biological activity. Furthermore, the labeled peptide exhibited saturation binding to EGF receptor-positive tumor cell lines which could be inhibited by presaturation of the cells with unlabeled, native EGF. Biodistribution in normal mice at 3 h postadministration showed rapid clearance with minimal retention of the label in sampled organs. We have now investigated the tumor localization properties in mice of this labeled peptide. Nude mice implanted with the EGF receptor-positive tumors A431 and LS-174T were administered labeled EGF and a labeled control peptide (BPTI, aprotinin). Tumor uptake at 12 h postadministration was 0.44% injected dose/g for EGF/g vs. 0.09 for the control. Pretreatment of tumored mice with unlabeled EGF blocked about half the tumor uptake. Animals were also administered an anti-EGF receptor antibody labeled with 99mTc via MAG3. Relative to the antibody, tumor-to-muscle ratios were improved from 6 to 15 and tumor-to-blood ratios from 0.4 to 7 with EGF. These favorable results along with documented evidence of overexpression of the EGF receptor in many human tumors suggest that 99mTc-EGF should be considered further for tumor detection.

Animals↗

PubMed1997

In vivo hybridization of technetium-99m-labeled peptide nucleic acid (PNA).

UNLABELLED: Hybridization of a radiolabeled single-stranded DNA oligonucleotide with its single-stranded complement in vivo has not yet been convincingly demonstrated. A contributing factor may be unfavorable in vivo properties of the phosphodiester and phosphorothioate DNAs. Peptide nucleic acid (PNA) oligomers have been reported to possess in vivo properties more suitable for radiopharmaceutical applications. METHODS: We have radiolabeled an amine-derivatized 15-base PNA oligomer with 99mTc through a modified MAG3 chelator. RESULTS: The ability of the PNA to hybridize in vitro with its complement appeared to be unimpaired after conjugation and radiolabeling. Size-exclusion, high-performance liquid chromatography (HPLC) analysis of 37 degrees C serum after 24 hr of incubation showed the radiolabel to be present predominately as labeled PNA with indications of labeled serum proteins and a low molecular weight catabolite. Whole-body clearance in mice was rapid, with 50% of the label eliminated in about 2 hr. After 2.5 hr, the highest uptake (kidneys) was only 1.5% of the injected dose/g; less than 0.07%/g was present in all sampled tissues at 24 hr. To evaluate in vivo hybridization, beads were implanted subcutaneously in both thighs of normal mice. In the left thigh only, the beads were conjugated with complementary single-stranded PNA. At 23 hr following intraperitoneal administration of the labeled PNA, the left/right thigh radioactivity ratio was 6:1. Whole-body images at this time showed only bladder, kidneys and the left thigh. CONCLUSION: Unlike the radiolabeled DNAs investigated in this laboratory, 99mTc-PNA displays stability and pharmacokinetic properties suitable for eventual use as radiopharmaceuticals.

Animals↗

PubMed1996

Technetium-99m antibodies labeled with MAG3 and SHNH: an in vitro and animal in vivo comparison.

The in vitro stability and animal pharmacokinetics of 99mTc bound to Sandoz and C110 IgG antibodies via a modified MAG3 has been compared with the hydrazino nicotinamide (SHNH) moiety as standard. For both antibodies, the stabilities of the label to challenge at up to 50:1 cysteine: IgG molar ratio were comparable, but at higher molar ratios, MAG3 showed greater instabilities. For the Sandoz antibody, size-exclusion HPLC analysis of 37 degrees C serum incubates and plasma samples from injected mice showed no clearly distinguishable differences. In the C110 case, some increased high molecular weight radioactivity was apparent with MAG3. Biodistributions in normal mice showed significant differences only in liver (Sandoz) and liver, spleen, intestines, stomach, and blood (C110), with SHNH usually providing higher levels. Thus, for two different antibodies and under the conditions of this study, the MAG3 chelator provided a 99mTc label with properties similar to that of SHNH moiety.

Animals↗

PubMed1996

Protein labelling via deoxyribonucleic acid hybridization.

A novel method of radiolabelling antibodies and other proteins is described. A small single-stranded DNA was covalently conjugated to an antibody and labelled by hybridization following the addition of the complementary single-stranded DNA labelled with technetium-99m (99Tc(m)) or indium-111 (111In). Antibody labelling efficiencies were 100% in about 1 h at room temperature with specific activities of up to 30 microCi micrograms-1 of IgG for 99Tc(m). Both diester and thioate DNAs were used. Both the diester- and thioate-labelled antibodies showed complete label stability in 37 degrees C saline. After 24 h in 37 degrees C serum, however, about 40% of the label in the case of the diester antibody was on low molecular weight species--probably labelled catabolites from nuclease degradation of the phosphodiester DNA. In contrast, the 99Tc(m) label on the thioate antibody was immediately and quantitatively bound to serum proteins--probably due to non-specific binding through the sulphur groups. Biodistribution studies in normal mice reflect these in vitro observations: 99Tc(m) on the diester antibody was rapidly cleared through the kidneys, probably as low molecular weight catabolite, while on the thioate antibody, the 99Tc(m) label was predominately deposited in the liver. In conclusion, by modifying with a single-stranded DNA, proteins may be readily labelled with a variety of radionuclides by DNA hybridization. The properties of the radiolabel are strongly influenced by the nature of the DNA.

Animals↗

PubMed1996

Comparative properties of a technetium-99m-labeled single-stranded natural DNA and a phosphorothioate derivative in vitro and in mice.

Oligonucleotides, particularly single stranded, may ultimately be of considerable use as radiopharmaceuticals. We have compared a synthetic 22-base single-stranded phosphodiester DNA with its phosphorothioate analog after both were radiolabeled with 99mTc via the hydrazino nicotinamide chelator. Whole body clearance of the label in mice was much slower when introduced on the phosphorothioate (30% vs. 75% clearance at 6 hr) because of immediate and persistent accumulation in liver (47% vs. 2% injected dose/g at 4 hr). The label in both cases was present in urine primarily on low molecular weight catabolites. High-performance liquid chromatography analysis of 37 degrees C serum incubates showed serum protein binding of 99mTc in both cases (about 100% bound at 24 hr) but to different proteins. Different behavior with respect to protein binding was also observed in the analysis of liver and kidney homogenates: the phosphodiester label was almost quantitatively converted to lower molecular weight catabolites after only 15 min, whereas the phosphorothioate label was primarily on proteins. The rapid digestion of the phosphodiester by nucleases was not observed, probably because protein binding of the labeled oligonucleotides stabilized against degradation. Thus the phosphodiester DNA may be the preferred 99mTc-labeled oligonucleotide in certain circumstances to avoid the high and persistent liver uptake observed with the phosphorothioate DNA.

Animals↗

PubMed1996

Imaging osteomyelitis with streptavidin and indium-111-labeled biotin.

UNLABELLED: Animal studies of infection imaging by a two-step protocol have shown that important improvements in target to nontarget ratios are possible. In this protocol, unlabeled streptavidin is administered and allowed sufficient time to accumulate in the lesion, probably by nonspecific processes, and to clear elsewhere. Thereafter, 111Inbiotin is administered. A fraction of the labeled biotin may be retained in the lesion because of biotin's high affinity for streptavidin while most of the activity is cleared through the kidneys. METHODS: Radioscintigraphy with unlabeled streptavidin followed with 111Inlabeled biotin was performed in 15 patients with chronic osteomyelitis. As controls, each patients received either 111In-labeled biotin without the preadministration of streptavidin or 111In-labeled nonspecific IgG. RESULTS: Regions of focal uptake were identified in all patients receiving streptavidin followed by radiolabeled biotin as early as 10 min postadministration of radioactivity, and retention of label was evident through 24 hr. Coincident regions of abnormal accumulation were apparent with 111In-IgG, but only in delayed images. Moreover, with 111In-biotin alone, without the preadministration of streptavidin, focal accumulations were detected in areas similar to that identified with the two-step protocol. Although, these observations were only in the earliest images. CONCLUSION: The results of this preliminary clinical investigation suggest that a two-step protocol with unlabeled streptavidin and radiolabeled biotin may be an alternative for the detection of infection.

Adult↗

PubMed1996

Investigations of directly labeling antibodies with rhenium-188.

Methods for labeling antibodies with 99mTc cannot be used without modification for radiorhenium despite the similar chemistries, in part because of a lower redox potential of rhenium and therefore a greater tendency to reoxidize. We have investigated conditions for directly labeling B72.3 IgG with 188Re via both mercaptoethanol and stannous ion antibody reduction. The reduced 188Re was stabilized for transchelation as the glucoheptonate complex and transchelated in the presence of excess stannous ion. End points were low "non-specific" binding (i.e. labeling in the absence of antibody reduction) and increased stability to cysteine challenge. By both methods, labeling efficiencies after about 15 minutes averaged 58.77% with as little as 4% non-specific binding. Specific activities of 15 muCi/microgram was achieved after 1.5 hours. By investigating labeling condition, it was possible to improve the stability of the label on stannous ion reduced antibody such that the in vitro and in vivo properties of 188Re were largely independent of labeling method. For example, losses of 188Re due to oxidation (16%) and to cysteine (7%) during 37 degrees C serum incubations for 24 hours were identical for both methods. Furthermore, after the administration to normal mice, whole body clearance and the accumulations of 188Re at 2.5 and 24 hours in blood and in most organs were also independent of labeling method. In conclusion, two different direct labeling methods provided a 188Re-labeled antibody with identical stabilities and with in vivo properties not greatly different from that seen for the same antibody radiolabeled directly with 99mTc.

Animals↗

PubMed1995

Influence of endogenous biotin on the biodistribution of labelled biotin derivatives in mice.

Previously, this laboratory reported that in mice pre-targeted with unlabelled streptavidin, the biodistribution of 111In administered on one biotin derivative (EB1) was superior to that of another derivative (DB2). In addition, a Scatchard analysis showed that the affinity constant of 111In-EB1 is lower by seven orders of magnitude from that of 111In-DB2. Therefore, this paper considers the role that endogenous biotin may play in these observations. Both 111In-labelled EB1 and DB2 were bound to streptavidin and incubated at 37 degrees C in mouse blood with increasing concentrations of d-biotin. As determined by Sephadex G-50 chromatography, only an 8-fold molar excess of d-biotin relative to labelled streptavidin was required to displace 90% of label in the case of EB1, whereas even a 20-fold molar excess provided no detectable displacement of DB2. That this displacement was also occurring in vivo was established in a mouse model bearing an infected thigh: increasing the serum biotin level (by intraperitoneal administration of d-biotin) had no effect on the biodistribution of 111In when administered on DB2; however, the target to non-target ratio decreased in the case of EB1. We have also observed that the biodistribution is no longer favourable when EB1 is administered radiolabelled with 99Tcm. When 111In was substituted with 99Tcm on EB1, chromatography of blood samples showed that similar displacement was occurring; however, in this case, the displaced label bound to serum proteins.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1995

Technetium-99m labeling of DNA oligonucleotides.

UNLABELLED: Single-stranded RNA and DNA oligonucleotides may be useful as radiopharmaceuticals for antisense and other in vivo applications if convenient methods for stably attaching radionuclides such as 99mTc can be developed. METHODS: To radiolabel DNA with 99mTc, we have used the hydrazino nicotinamide (SHNH) moiety developed elsewhere. The diethylenetriaminepentacetic acid (DTPA) chelate was used to label DNA with 111In for comparison. Complementary 22-base, single-stranded oligonucleotides were obtained, each with a primary amine attached to either 3' or 5' end with a biotin moiety on the opposite end. The DNA was conjugated with SHNH by a N-hydroxysuccinimide derivative with DTPA by the cyclic anhydride. RESULTS: Reversed-phase HPLC analysis showed that essentially complete conjugation was achieved in both cases. The purified SHNH-DNA was radiolabeled with 99mTc by transchelation from glucoheptonate at labeling efficiencies of up to 60% and DTPA-DNA with 111In acetate at up to 100% efficiency. After labeling, the ability of the DNAs to bind to streptavidin through the biotin moieties and to hybridize with their complementary DNA in saline was retained for both radiolabels as determined by size-exclusion HPLC analysis. HPLC radiochromatograms of serum incubates showed a shift to 99mTc, but not 111In, to a high molecular weight, strongly suggesting serum protein binding in the former case only. Low-molecular weight degradation products were seen with 111In, but not with 99mTc and may be related to the use of phosphodiester-linked oligonucleotides. As a further measure of label stability, the DNAS were bound to streptavidin-conjugated magnetic beads and incubated in fresh 37 degrees C human serum. Less than 4% of 99mTc and 14% of 111In was lost in 24 hr. CONCLUSION: Amino-modified, single-stranded DNA can be stably radiolabeled with 99mTc by the SHNH moiety without loss of function.

Animals↗

PubMed1994

Can a cysteine challenge assay predict the in vivo behavior of 99mTc-labeled antibodies?

Recent investigations have shown that transchelation to cysteine in a principal mode of in vivo instability of 99mTc-labeled antibodies. In this investigation, a cysteine challenge assay was used to measure the in vitro instability of 99mTc directly labeled to two IgG antibodies (B72.3 and C110) via two established direct labeling methods employing mercaptoethanol and stannous ion for antibody reduction and by a novel method using glutathione for this purpose. For both antibodies, the greatest instability to cysteine occurred with stannous ion reduction. The stability of glutathione-reduced B72.3 was indistinguishable from mercaptoethanol-reduced B72.3 whereas glutathione-reduced C110 showed stability roughly intermediate between that of the other reducing agents for this antibody. Results obtained in normal mice were in the direction predicted by the assay: for both antibodies, urinary clearance of 99mTc was fastest in mice receiving antibodies labeled via stannous ion reduction, presumably because of the increased transchelation of label to cysteine in vivo. Urinary clearance was slower and identical in mice receiving B72.3 labeled via glutathione or mercaptoethanol whereas clearance in the case of glutathione-reduced C110 was intermediate between that of the other two reducing agents. At both time points, higher radioactivity levels were observed in kidneys and lower levels in blood and most other tissues for both antibodies in the case of stannous ion reduction as expected for the label of greatest instability. In the B72.3 case, with only one exception, tissue and blood levels following administration of glutathione-reduced antibody were indistinguishable from that following administration of mercaptoethanol-reduced antibody. In the C110 case, significant differences in activity levels were observed in several tissues between glutathione- and mercaptoethanol-reduced antibodies. In conclusion, the relative in vivo behaviour of 99mTc when administered to mice while labeled to two IgG antibodies were successfully predicted based on the results of an in vitro cysteine challenge assay.

Animals↗

PubMed1994

Investigations of ascorbate for direct labeling of antibodies with technetium-99m.

UNLABELLED: Recently, a method for the direct labeling of antibodies with 99mTc was described in which sulfhydryls were reportedly generated by reduction of antibody disulfides with ascorbic acid. Thereafter, these proteins may be labeled at high efficiency with 99mTc following reduction of pertechnetate with dithionite. This investigation was initially conducted to evaluate the mechanism of the increased stability towards cysteine challenge reported for the label and subsequently to determine the role of ascorbate in the labeling process. METHODS: It was possible to reproduce the reported high labeling efficiencies by increasing the dithionite concentration fivefold, presumably because of variabilities among lots of commercial sodium dithionite. RESULTS: Despite success in labeling, it was not possible to confirm that antibody reduction followed the treatment with ascorbate. Using both Ellman's reagent and 2,2' dithiodipyridine as indicators, we were unable to detect sulfhydryls on one IgG antibody treated at ten times the suggested ascorbate-to-antibody molar ratio. It was estimated that the number of sulfhydryls generated could not have been more than 1% (dithiodipyridine) to 2% (Ellman's). Furthermore, radiolabeling efficiencies for two IgG antibodies and stabilities of the label to cysteine challenge were unchanged when the ascorbate was eliminated. The number of sulfhydryls generated by treatment of the antibody with dithionite at 1-2 times the concentration required for adequate labeling was about 1% (dithiodipyridine) to 5% (Ellman's). CONCLUSION: For the conditions of this investigation and for the antibodies employed, ascorbate apparently played no more than a minor role at best in the labeling process. If antibody reduction occurred, this most likely was a result of residual dithionite presented to the protein along with the reduced 99mTc.

Antibodies↗

PubMed1993

Improved tumor localization with (strept)avidin and labeled biotin as a substitute for antibody.

Because of its short physical half life, the use of anti-tumor antibodies radiolabeled with 99mTc has necessitated early (i.e. 2-6 h post-administration) imaging. It is possible that at these early times localization of antibodies in certain tumors may be largely due to non-specific processes. If so, other proteins or agents may be preferred for early imaging of solid tumors. We have investigated tumor localization with labeled biotin administered subsequent to unlabeled and unconjugated streptavidin. Nude mice bearing anti-CEA tumors (LS174T) received 10 micrograms of 111In-labeled anti-CEA antibody (C110) or 111In-labeled streptavidin with sacrifice 5 h later. In an examination of pretargeting, other animals received 50 micrograms of unlabeled streptavidin followed 3 h later with 1 micrograms of 111In-labeled biotin (EB1) and sacrifice 2 h later. The biodistribution of labeled streptavidin was similar to that of labeled specific antibody except for lower blood and higher kidney levels. Tumor levels were also lower with labeled streptavidin but, because of still lower levels in liver and blood, the tumor/normal tissue ratios were improved. When unlabeled streptavidin was administered and followed by labeled biotin (pretargeting), tumor levels were further reduced modestly; however, normal tissue levels were greatly reduced such that the tumor/blood and tumor/liver ratios were 10.6 and 2.2 vs 1.5 and 0.5 for the specific antibody. Improvements were seen in all tissues sampled with the exception of kidney and muscle. A further control of labeled biotin alone (without the preinjection of streptavidin) showed minimal accumulations in all tissues with the exception of kidneys.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1993

Pharmacokinetics of the FO23C5 anti-CEA antibody fragment labelled with 99Tcm and 111In: a comparison in patients.

The FO23C5 anti-carcinoembryonic antigen (CEA) F(ab')2 antibody was radiolabelled with 111In via diethylenetriaminepentaacetic acid (DTPA) and directly with 99Tcm by stannous ion and mercaptoethanol antibody reduction to compare the pharmacokinetics of these three agents. Four patients received 15 mCi 99Tcm-Fab' 1 week before receiving 1 mCi 111In-F(ab')2. Five additional patients received only the 99Tcm-Fab'. Radiochromatograms by high-performance liquid chromatographic (HPLC) analysis of serum and urine samples from patients receiving 111In were typical of those observed by us previously in connection with other antibodies. The identical analyses of samples from patients receiving 99Tcm showed no differences with the method of reduction and more complex radiochromatograms. In addition to peaks due to a mixture of labelled F(ab')2 and Fab' fragments and, occasionally, immune complexes, there were several peaks due to labelled cysteine and other small labelled species present in both serum and urine. The biodistribution of 99Tcm was as expected for a labelled Fab' fragment: relative to 111In, 99Tcm cleared rapidly from circulation and into kidneys and urine. Liver levels of 111In and 99Tcm were surprisingly similar at 1 day (12 versus 9% ID) although initial 111In levels were lower and increased while 99Tcm levels were higher and decreased. Spleen levels were also similar. In 4/9 patients receiving 99Tcm, hepatobiliary clearance was observed at levels which could confuse interpretation whereas this mode of clearance was observed in only 1/4 patients receiving 111In. Image quality was superior with 111In versus 99Tcm at 1 day postadministration as judged by counting rates and background activity whereas the opposite was true at 2-3 h postadministration.

Adult↗

PubMed1993

Directly and indirectly technetium-99m-labeled antibodies--a comparison of in vitro and animal in vivo properties.

To investigate the in vivo and in vitro properties of 99mTc when labeled to antibodies via one direct and one indirect method, the B72.3 and C110 IgG antibodies were radiolabeled directly via stannous ion reduction and indirectly via the hydrazino nicotinamide chelator and compared in vitro and in vivo. Antibody avidity (but not immunoreactive fraction) appeared to be independent of labeling methods for both antibodies. Following stannous ion reduction, antibodies were fragmented by denaturing SDS PAGE although only slight evidence of fragmentation was found in vivo. The direct label was instable to transchelation to cysteine and glutathione in vitro and in vivo. Following intravenous administration, urinary excretion of activity was threefold greater for the direct label and was almost exclusively labeled cysteine and glutathione. Significant differences in the biodistribution of 99mTc were also observed: liver levels were lower, kidney levels were higher and clearance of label from blood and tissues was faster for the direct label. At Day 1, tumor accumulation was threefold lower for the direct label although most normal tissues were also lower. In conclusion, when labeled to two antibodies by one direct method, 99mTc is unstable towards transchelation relative to one indirect method. These relative instabilities greatly influenced the biodistributions in mice and may influence the quality of images obtained in patients.

Animals↗

PubMed1992

The stability of 99Tcm directly labelled to an Fab' antibody via stannous ion and mercaptoethanol reduction.

The anti-CEA FO23C5 F(ab')2 antibody was directly radiolabelled with 99mTcm by two methods (stannous ion and mercaptoethanol reduction) and compared in vitro and in vivo for label stability. By both methods, reduction of the F(ab')2 fragment produced primarily Fab' fragments. By both methods, the label was stable to 99Tcm-pertechnetate formation in vitro. Analysis by high performance liquid chromatography (HPLC) of serum, urine, kidney and liver homogenates from mice injected with 99Tcm-antibodies by both methods consistently showed a prominent radiolabelled peak with an estimated molecular weight of about 300 daltons. An identical peak was observed in the analysis of patient samples in a related investigation from this laboratory. Cysteine was radiolabelled with reduced 99Tcm and analysed by HPLC and thin layer chromatography (TLC); one of the 99Tcm-cysteine species so produced showed the same chromatographic behaviour as that of the 300 dalton species. In conclusion, the FO23C5 and other antibodies are stably labelled with 99Tcm via either stannous ion or mercaptoethanol reduction. In mice and in patients, the labelled proteins are either catabolized or, more likely, the 99Tcm label is transchelated such that the label is present on several low molecular weight species, the most prominent of which is postulated to be 99Tcm-cysteine.

Animals↗

PubMed1992

Localization of infection using streptavidin and biotin: an alternative to nonspecific polyclonal immunoglobulin.

Since favorable images of infection are obtained with radio-labeled nonspecific IgG, streptavidin has been considered as an alternative protein in this investigation. The advantage of streptavidin is that once localized it may be targeted with radiolabeled biotin. Studies were conducted in a mouse model with an Escherichia coli infection in one thigh. Indium-111-labeled streptavidin showed equivalent localization to the infection as that obtained with 111In-labeled polyclonal nonspecific IgG, however blood levels with streptavidin were lower at all time points; consequently, target-to-blood ratios were improved. Pretargeting with unlabeled streptavidin followed 3 hr later with 111In-labeled biotin showed equivalent localization in the target and reduced activity in all organs sampled. As such, infected thigh-to-normal thigh ratios were improved 3-fold for pretargeting versus either labeled IgG or streptavidin. Improvements in infected thigh-to-liver and blood ratios were greater than 8-fold. Only in the case of kidneys was the ratio unimproved. In conclusion, we have shown that by preadministration of unlabeled streptavidin followed by labeled biotin, infectious lesions in a mouse model may be imaged earlier with lower background levels relative to the administration of labeled nonspecific IgG.

Animals↗