 |
||||
| 2007 JASN IMPACT FACTOR 7.111 | HOME AUTHOR INFO EDITORIAL BOARD SUBSCRIBE FEEDBACK ALERTS HELP | |||
| CURRENT ISSUE | ARCHIVES | JASN Express | ONLINE SUBMISSION | |
|
||||
| 2007 JASN IMPACT FACTOR 7.111 | HOME AUTHOR INFO EDITORIAL BOARD SUBSCRIBE FEEDBACK ALERTS HELP | |||
| CURRENT ISSUE | ARCHIVES | JASN Express | ONLINE SUBMISSION | |
*
Department of Medicine, Division of Nephrology and Hypertension, State
University of New York at Stony Brook, Stony Brook, New York
Department of Physiology, Division of Nephrology and Hypertension, State
University of New York at Stony Brook, Stony Brook, New York
Division of Cardiology, State University of New York at Stony Brook, Stony
Brook, New York
Ciphergen Biosystems, Inc., Palo Alto, California.
Correspondence to Dr. Michael S. Golgorsky, Department of Medicine, Division of Nephrology and Hypertension, State University of New York at Stony Brook, Health Science Center T15-020, Stony Brook, NY 11794-8152. Phone: 631-444-1617; Fax: 631-444-6174; E-mail: mgoligorsky{at}mail.som.sunysb.edu
 |
Abstract |
|---|
 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
|
Introduction |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
The recent development of surface-enhanced laser desorption/ionization (SELDI)-time of flight (TOF) technology, based on improved methods for the chemical preparation of absorptive surfaces and their use for solid-state mass spectrometry (5), allows high-throughput protein analysis of crude biologic samples. Key components of this technology, in addition to the mass spectrometer, are ProteinChip arrays that contain 2-mm-diameter adsorptive target spots. Each of these targets represents either a chemical (e.g., ionic, hydrophobic, or hydrophilic) or immunoabsorptive (antibody) surface designed to capture proteins of interest. After removal of unbound proteins and interfering substances, the molecular masses of the proteins retained on the ProteinChip can be determined by TOF analysis. The potential advantages of SELDI ProteinChip technology include rapidity and reproducibility in the screening of protein expression profiles known as "phenomic fingerprints." In general, this technology can be used to provide phenomic fingerprints of complex protein mixtures; however, there are no published data on the use of this technique in studies of normal and pathologic urinary protein profiles. This work summarizes our investigation designed to test the utility (accuracy, reproducibility, and sensitivity) of this technique in studies of urinary protein composition and to exemplify its use with a study of the changes in urine composition associated with radiocontrast medium administration. This latter condition was selected for two main reasons, i.e., the large number of patients who undergo cardiac catheterization and the poor predictability of its renal complications.
The incidence of acute renal failure triggered by the intravenous administration of radiocontrast media for diagnostic purpose ranges from 1.2 to 100%, depending on the number of risk factors (6,7,8). Despite the routine precautionary measures taken for each patient, radiocontrast medium-induced nephropathy remains a serious risk for procedures (such as cardiac catheterization) performed in already-compromised hosts. This condition is especially dangerous because overt signs of such complications are detectable 24 to 48 h after the procedure, when patients have been discharged or have been scheduled for or even undergone cardiac surgery. Early knowledge of impending renal complications could significantly reduce the morbidity associated with this diagnostic procedure. The existing markers of developing radiocontrast medium-induced nephropathy, based on measurements of enzymuria, are non-specific, and their detection remains technically cumbersome (2). SELDI-TOF-generated data on urinary protein compositions after radiocontrast medium administration are summarized in this report.
|
Materials and Methods |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
The ProteinChip system was calibrated with bovine superoxide dismutase and
bovine ß-lactoglobulin A (both from Sigma Chemical Co., St. Louis, MO).
Identification of 
-defensin-2 [human neutrophil peptide-2 (HNP-2)] in
urine samples, on the basis of mass, was performed using the SWISS-PROT
protein database, allowing 0.1% error in observed mass. To confirm the
identity, the mass of synthetic human HNP-2 (Sigma) was compared with findings
for the urine sample. To assess the sensitivity of the ProteinChip system,
purified HNP-2 was diluted from 10 fmol to 0.1 fmol and the peak amplitudes
were determined. Peaks with amplitudes at least 3 times greater than the
average background noise level were considered meaningful. The reproducibility
was tested by depositing different aliquots of the same urine sample on six
different spots of the ProteinChip array.
Identification of ß2-Microglobulin
To confirm the identity of ß2-microglobulin (ß2M), the
masses of purified ß2M from human urine (Calbiochem, San Diego, CA) and
immunoprecipitated ß2M from the samples were compared with findings for
the original urine samples. For immunoprecipitation of ß2M, urine samples
were incubated for 2 h at 4°C, with agitation, either with anti-ß2M
antibodies (Sigma) or with irrelevant polyclonal goat anti-mouse antibodies
(Jackson ImmunoResearch Laboratories, West Grove, PA) as a negative control.
After incubation for 1 h at 4°C, with agitation, with protein A/G beads
(Protein A/G PLUS agarose; Santa Cruz Biotechnologies, Santa Cruz, CA) in a
1:1 suspension in phosphate-buffered saline, the samples were centrifuged at
10,000 x g for 5 min at 4°C. The beads were washed twice
with washing buffer I (10 mM Tris-HCl, pH 7.4, 150 mM NaCl, 1 mM
ethylenediaminetetraacetate, 1% Triton X-100, 1% Nonidet P-40, 1 µg/ml
leupeptin, 1 µg/ml aprotinin, 10 µg/ml trypsin inhibitor, 0.5 mM
phenylmethylsulfonyl fluoride, 1 mM sodium orthovanadate) and once with
washing buffer II (10 mM Tris-HCl, pH 7.4, 1 mM ethylenediaminetetraacetate).
After final centrifugation, the pellets were reconstituted in 20 µl of
water.
To dissociate antigen-antibody complexes from the protein A/G beads, the samples containing the beads were vortex-mixed for 2 min at 37°C and then cooled on ice. After addition of elution buffer (ImmunoPure Gentle Ag/Ab Elution Buffer; Pierce, Rockford, IL) to each of the samples and vortex-mixing for 15 min at room temperature, the samples were centrifuged at 16,000 x g for 10 min. Guanidine (8 M in 0.25% trifluoroacetic acid) was added to the pellet, and the mixture was vortex-mixed for 15 min at room temperature and centrifuged at 16,000 x g for 2 min. Solid-phase extraction was performed using ZipTipC18 cartridges (Millipore Corp., Bedford, MA), according to the instructions provided by the manufacturer, except that proteins were eluted with 2,2,2-trifluoroethanol (Aldrich, Milwaukee, WI).
Animal Studies
All experiments were performed in accordance with the NIH Guide for the
Care and Use of Laboratory Animals
(9). Twenty male Sprague-Dawley
rats (270 to 290 g) were maintained on standard rat chow, with water available
ad libitum and with an automatically controlled 12-h light/dark
cycle, in the accredited Department of Laboratory Animal Research at the State
University of New York at Stony Brook. The animals were placed in individual
metabolic cages, and baseline 24-h urine samples were collected after a 1-d
adaptation period. The rats were divided into four experimental groups of five
animals each. The animals in groups 1 and 3 received hypertonic saline
solution (690 mOsm) intravenously and were used as control animals. The rats
in groups 2 and 4 received intravenous injections of 1 ml (1.25 g/kg body wt)
of the nonionic, low-osmolar (695 mOsm), radiocontrast agent ioxilan
(Oxilan-350; Cook Imaging Corp., Bloomington, IN), which was first described
in 1988 (10). One-half of the
rats (groups 3 and 4) were pretreated with
NG-nitro-L-arginine methyl ester (L-NAME) (50 mg/L in
water) to mimic the endothelial dysfunction observed in patients with
cardiovascular disease (modified from references
11 and
12). Twenty-four-hour urine
samples were collected for 2 d consecutively. Serum and urine creatinine
levels were measured by colorimetric assay (Beckman Synchron CX and kit
443340); blood urea nitrogen (BUN) levels were measured using the enzymatic
conductivity rate method (Beckman Synchron CX and kit 443350).
Clinical Studies
Twenty-five patients admitted to the University Hospital of the State
University of New York at Stony Brook for diagnostic or therapeutic cardiac
catheterization were recruited to participate in the study. Patients who had
undergone any invasive procedure 2 wk before admission and those with severe
nephrotic syndrome, active lupus nephritis, genitourinary tract infection, or
liver failure were excluded from the study. Three freshly voided urine samples
were studied; the first was obtained before cardiac catheterization, the
second immediately after it, and the third approximately 6 to 12 h after the
procedure. In the course of routine follow-up monitoring, serum creatinine and
BUN levels were measured 48 h after the procedure. The patients were divided
into two groups; one group consisted of patients with normal renal function,
as indicated by plasma creatinine levels (n = 20), and the other
group consisted of patients with deteriorations in renal function (n
= 5). Ioxilan was administered as a radiocontrast agent to all patients
studied. The study was approved by the Committee on Research Involving Human
Subjects of the State University of New York at Stony Brook. All patients
signed an informed consent form for participation.
Sample Preparation
Urine samples were immediately centrifuged at 3000 x g, the
sediments were analyzed microscopically, and the supernatants were stored
frozen at -80°C until further analysis. Supernatants were slowly thawed on
ice and either filtered at 7500 x g for 20 min at 4°C
through a Microcon centrifugal filter device with a 50-kD cut-off (Millipore)
or analyzed directly. The retentates were filtered three times and each time
reconstituted with deionized water.
Statistical Analyses
Results are reported as mean ± SEM. Comparisons between control
samples and each of the treatment groups were made using t tests.
Analyses of differences in multiple groups were performed using one-way ANOVA.
A P value of <0.05 was considered to indicate a statistically
significant difference.
|
Results |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
|
Figure 2 displays protein
peaks in the range between 3000 and 4000 D. In
Figure 2 (upper), the test
urine sample demonstrates three unknown peaks, with apparent molecular masses
of 3370, 3441, and 3484 D. Using the SWISS-PROT protein database, peaks with
these masses were tentatively identified as HNP-2 (calculated masses, 3371.0
D), HNP-1 (calculated mass, 3442.1 D), and HNP-3 (calculated mass, 3486.2 D).
Human -defensins-1 to -3 (also termed HNP-1 to -3, respectively) are a
group of small cationic peptides (average mass, 3.3 to 3.5 kD) that are
derived from neutrophils and exhibit broad-spectrum antimicrobial activity
(13,14).
They are almost identical in sequence. HNP-1 and -3 are 30 amino acids long
and differ only at the first amino acid, which is alanine in HNP-1 and
aspartate in HNP-3. HNP-2 is only 29 amino acids in length
(15). Application of purified
HNP-2 peptide to the target, followed by SELDI-TOF analysis, revealed a peak
at 3369.6 D (<0.05% difference from the mass measured in the complex
biologic sample), thus confirming the presence of HNP-2 in the test urine
sample.
|
The sensitivity of SELDI-TOF is illustrated in
Figure 3; sensitivity was
assessed by titration of HNP-2 from 10 fmol down to 0.1 fmol. A level of 0.1
fmol of HNP-2 yielded a peak at 3370 D that was significantly greater than the
background noise, and this level was defined as the detection limit in this
setting. The test urine sample used to determine the reproducibility
demonstrated, among other proteins, the triple peak that is characteristic of
-defensins (Figure 4).
The peak representing HNP-2 demonstrated an average mass of 3372.2 D (SD,
±0.1; calculated mass, 3371.0 D). The protein profiles of the test
urine sample preabsorbed on six target spots of the ProteinChip array were
nearly identical. Collectively, these data demonstrate that SELDI-TOF is a
highly sensitive, reproducible, accurate technique that allows the detection
of minute protein fractions in biologic samples as complex as urine. These
findings permitted the use of SELDI-TOF to study urinary protein composition
of experimental animals and patients receiving radiocontrast medium.
|
|
Effects of Radiocontrast Medium in Experimental Animals
The four groups of rats received the nonionic radiocontrast medium ioxilan
or vehicle. The rate of urine output was unaffected by this treatment
(Figure 5C). There was a
marginal decrease in creatinine clearance in animals that received either
ioxilan or L-NAME alone, from 3.63 ml/h to 2.79 and 2.90 ml/min, respectively.
In contrast, animals that received both ioxilan and L-NAME exhibited a
significant reduction in creatinine clearance to 2.38 ml/min (P <
0.05, compared with control), as summarized in
Figure 5B.
|
The protein profiles of the urine samples from control rats that were studied using SELDI-TOF indicated that the most prominent peaks occurred at 9.9, 18.7, 21.0, 33.3, and 66.3 kD (Figure 5A). For animals pretreated with L-NAME, there were only minor changes in the 21.0-, 33.3-, and 66.3-kD proteins. For rats given infusions of ioxilan, amounts of a protein with a molecular mass of 21 kD were decreased. In contrast to these relatively minor changes, a dramatic reduction or disappearance of all five protein peaks was observed in the urine of rats treated with both ioxilan and L-NAME, which was the group characterized by reduced creatinine clearance.
Effects of Radiocontrast Medium in Patients
Plasma creatinine and BUN levels in patients with normal kidney function
exhibited no significant differences before and after cardiac catheterization
(creatinine, 0.9 ± 0.06 versus 0.8 ± 0.05 mg/dl,
P = 0.455; BUN, 16.3 ± 1.3 versus 15.6 ± 1.28
mg/dl, P = 0.404). In patients with impaired renal function, plasma
creatinine and BUN levels were elevated before the catheterization and plasma
creatinine levels exhibited a trend toward further increase after the
procedure (creatinine, 1.9 ± 0.24 versus 2.2 ± 0.37
mg/dl, P = 0.152; BUN, 41.4 ± 13.0 versus 40.2
± 13.7 mg/dl, P = 0.235). Urinalyses and sediment analyses
revealed no major interval changes in the urine samples after cardiac
catheterization, compared with the initial samples.
The screening of patients with unimpaired renal function and without clinically detectable proteinuria demonstrated several prominent peaks detected by SELDI-TOF analysis, which represented proteins with molecular masses of 4.87, 9.75, 11.75, 23.5, 33.3, and 66.4 kD (Figure 6). Urine samples obtained after the administration of radiocontrast medium, i.e., immediately after the completion of cardiac catheterization, displayed changes in the abundance of normally detectable proteins; increases in proteins with molecular masses of 4.87, 9.75, and 11.75 kD were accompanied by the almost complete disappearance of a peak at 66.4 kD. Six to 12 h after the procedure, there was a strong trend toward the recovery of the precatheterization urinary protein pattern in patients with normal renal function who did not develop renal complications (as judged by plasma creatinine levels).
|
A strikingly different pattern of urinary proteins was observed for five patients with mild chronic renal insufficiency (Figure 7). Proteins with apparent molecular masses of 9.75, 11.75, 23.5, 33.3, and 66.4 kD, as well as several higher-molecular mass species of 79.3, 90.3, 133, and 199.2 kD (not shown), were detected using SELDI-TOF analysis. After cardiac catheterization and infusion of radiocontrast medium, urine samples exhibited the appearance of low-molecular mass peaks at 4.87 and 7.8 kD, increases in peaks at 9.75, 11.75, and 23.5 kD, and suppression of peaks at 33.3, 66.4, and 199.2 kD, which was not reversible within the first 6 to 12 h after the procedure. The 9.75-, 11.75-, and 23.5-kD peaks were increased even further after 6 to 12 h, as opposed to the patterns observed for patients with normal renal function, which tended to recover the baseline protein profile (Figure 6).
|
Because the 66.6-kD protein peak represented albumin and exhibited dynamic changes indicative of its perfusion-dependent excretory changes, we next used the amplitude of this peak as a common denominator for studies of the relative excretion of other proteins. The calculated ratio of the amplitude of the 9.75-kD or 11.75-kD peak to that of the 66.6-kD protein peak revealed marked amplification of differences for each of these peaks separately (Figure 8). There was a 10-fold increase in both ratios for patients with normal renal function immediately after cardiac catheterization, with a return to baseline values 6 to 12 h after the procedure. In contrast, for patients with impaired renal function, the 9.75 kD/66.6 kD ratio, which was initially <0.1, increased 60- and 70-fold immediately and 6 to 12 h after catheterization, respectively. The 11.75 kD/66.6 kD ratio also increased approximately 15-fold immediately after the procedure and remained elevated 6 to 12 h after the administration of contrast medium.
|
The 11.75-kD protein in the urine exhibited the same mass and pattern as purified ß2M from human urine (Figure 9A). Furthermore, the same peak could be observed in the immunoprecipitate but not the immunodepleted supernatant of samples incubated with specific anti-ß2M antibodies (Figure 9B). In contrast, urine samples incubated with irrelevant goat anti-rabbit antibodies as a negative control demonstrated no peak in the immunoprecipitate but an 11.744-kD peak in the supernatant (Figure 9C). The 11.75-kD peak thus can be safely identified as ß2M.
|
|
Discussion |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
The radiocontrast medium used in this study, ioxilan, is a modern,
nonionic, low-osmolar agent known to be stable, water-soluble, and well
tolerated (10), with
reportedly minimal effects on systemic and renal hemodynamics
(16,17,18).
Urine profiles, including albumin, glucose, lactate dehydrogenase,
N-acetyl-ß-D-glucosaminidase, and 
-glutamyltransferase
measurements, obtained for rabbits after administration of ioxilan
demonstrated no significant changes
(19). In experimental rats,
however, increases in albumin, lactate dehydrogenase, and
-glutamyltransferase excretion were reported
(20). In view of the reported
minimal nephrotoxicity of ioxilan, the changes in urinary protein composition
observed for both patients and experimental animals attest to the high
sensitivity of SELDI-TOF in detecting trace amounts of proteins, which are
usually undetectable by other techniques. However, dramatic changes in urinary
protein composition were detected mainly in patients and experimental animals
with pre-existing renal dysfunction. In the animal model that mimics the
endothelial dysfunction observed in patients with cardiovascular disease
(inhibition of endothelial nitric oxide synthase with L-NAME), there was a
significant reduction in creatinine clearance after the injection of ioxilan,
compared with control findings. This might reflect the limited nephrotoxicity
of radiocontrast agents themselves, and it is consistent with previously
reported observations
(11,12).
Concurrent with these functional abnormalities, SELDI-TOF analysis revealed a
different urinary protein pattern; all protein components observed in normal
urine were decreased or nearly absent
(Figure 5A). These findings
probably reflect the cumulative effects of contrast agent-induced
nephrotoxicity and vascular dysfunction, leading to reductions in GFR and
protein excretion. Even more dramatic changes were observed in human patients
with renal dysfunction (see above).
Using the SELDI-TOF technique, it has become possible to detect previously unrecognized proteins in urine obtained from patients with otherwise normal kidney function and no history of renal diseases. Specifically, we demonstrated that, in uncomplicated cases of radiocontrast medium infusion during cardiac catheterization, proteins with a broad range of molecular masses either appear or disappear from the urine, with the tendency to recover after 6 to 12 h (Figure 6). These might represent a previously unrecognized response of the kidney to the administration of radiocontrast medium. The increase in low-molecular mass proteins (9.75 and 11.75 kD) and the decrease in higher-molecular mass proteins (66.6-kD albumin) after the administration of radiocontrast medium can be attributed to combined glomerular and tubular dysfunction. Although it was expected that the repertoire and abundance of proteins in the urine of patients with known renal diseases would be more pronounced, SELDI-TOF analysis revealed a distinctive protein pattern and a lack of reversibility for changes in several protein components that appeared in the urine after cardiac catheterization. Surprisingly, there was no clear-cut correspondence of the urinary protein profiles in the experimental animals and the patients receiving ioxilan. However, possible differences in the molecular masses of proteins, attributable to different post-translational modifications or species-specific sequence variations, may be responsible for the observed lack of similarities between experimental animals and human subjects.
The analysis of SELDI-TOF protein peaks revealed substantial differences in the dynamics of protein patterns in the urine of patients with or without pre-existing renal insufficiency, as summarized in Figure 8. The latter group demonstrated a lack of reversibility in the depressed excretion of 66.6-kD albumin and increased excretion of proteins with molecular masses of 9.75 and 11.75 kD. The fact that these peptides exhibited changes in the opposite direction, compared with changes in albumin, argues against the possibility that changes in GFR are entirely responsible for the observed protein pattern dynamics. Although this could be true for albumin excretion, increased levels of proteins with molecular masses of 9.75 and 11.75 kD would be inconsistent with a causal role for renal hemodynamics. The observed reciprocal changes in proteins with molecular masses of 9.75 or 11.75 and 66.6 kD could be further exploited as a potentially useful index, i.e., 9.75 kD/66.6 kD or 11.75 kD/66.6 kD ratio. As demonstrated in Figure 8, these ratios are dramatically increased immediately and 6 to 12 h after cardiac catheterization in patients with impaired renal function, as opposed to patients with no renal complications. In future studies, we shall assess the utility of these indices in predicting impending contrast agent-induced nephropathy.
There is an urgent need for such early markers, because the frequency of renal complications resulting from radiocontrast medium administration remains high and therapeutic interventions to treat such complications are quite limited. Our search of protein databases identified several plausible candidates that were revealed using SELDI-TOF analysis. Species of 9.75 kD are most probably heparin-binding epidermal growth factor-like growth factor (9.729 kD; species of 4.87 kD are the same molecules but doubly charged). Proteins of 11.75 kD most likely represent ß2M (11.731 kD), as shown in Figure 9. The peak in the urine samples from patients with impaired renal function demonstrates the same mass and pattern as purified ß2M from human urine (Figure 9A). Furthermore, the same peak can be observed in the immunoprecipitate but not the immunodepleted supernatant of samples incubated with specific anti-ß2M antibodies (Figure 9B), as opposed to urine samples incubated with irrelevant goat anti-rabbit antibodies (Figure 9C). This is evidence that the protein represented by the peak with a molecular mass of 11.74 kD bound specifically to anti-ß2M antibodies and can thus be identified as ß2M. The candidate marker with an apparent molecular mass of 23.5 kD is most likely cathepsin O (23.460 kD). There is convincing evidence that the 66.4-kD (and doubly charged 33.3-kD) peaks observed in urine from rats and patients represent albumin (66.480 kD), especially from experiments in which the samples were mixed with an albumin standard (data not shown), although there is not yet an explanation for why albumin excretion decreases after the administration of radiocontrast medium. There are data to support heparin-binding epidermal growth factor-like growth factor playing an important role in the pathophysiologic processes of acute renal injury (21,22,23), making it a potentially meaningful marker. Different investigators have used ß2M as a marker of nephrotoxicity (24,25), with controversial results (26). However, there are no published data on the role of cathepsins as markers of any nephropathy. Definitive proof can be obtained through the isolation and microsequencing of these proteins. At that stage, it will be possible to perform a complete analysis of samples from patients receiving radiocontrast medium, in the search for candidate markers that are predictive of impending radiocontrast agent-induced nephropathy.
The proteins detected in the urine fall into several categories (4), as follows: (1) proteins that are also present in plasma and appear in the urine either intact or as fragments (>31 proteins) (27,28); (2) proteins that are produced by the kidney (approximately 30 different enzymes) or lower urinary tract (12 derived from the bladder, ureters, urethra, and/or prostate gland); (3) proteins that leak into the circulation, and eventually into the urine, from other organs, i.e., liver-, testicle-, or skeletal muscle/myocardium-specific proteins; (4) hormones or other signaling molecules (in the case of human chorionic gonadotropin, diagnosis of pregnancy has been made simple) (1); and (5) tumor-associated, bacterial, or viral products. These theoretical considerations should underscore the applicability of SELDI-TOF uroscopy (in conjunction with the analysis of serum) to a broader range of clinical problems.
In conclusion, the data presented here demonstrate the applicability of SELDI-TOF for the detection of unique proteins in the urine and the potential of this technique for the identification of marker proteins to aid in the prediction of impending renal complications of radiocontrast agent administration during procedures such as cardiac catheterization. As a proof of principle, we have identified one of the peaks in the SELDI protein profiles, i.e., that at 11.75 kD, as ß2M. Application of the SELDI-TOF technique to studies of urinary protein composition has the potential to revolutionize this field, making mass spectrometric uroscopy an important predictive and diagnostic tool.
|
Acknowledgments |
|---|
|
References |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
This article has been cited by other articles:
 |
|
 |
|
  K. J. A. Vanhoutte, C. Laarakkers, E. Marchiori, P. Pickkers, J. F. M. Wetzels, J. L. Willems, L. P. van den Heuvel, F. G. M. Russel, and R. Masereeuw Biomarker discovery with SELDI-TOF MS in human urine associated with early renal injury: evaluation with computational analytical tools Nephrol. Dial. Transplant., October 1, 2007; 22(10): 2932 - 2943. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. M. Wilson, E. Kimura, R. K. Harada, N. Nair, B. Narasimhan, X.-Y. Meng, F. Zhang, K. R. Beck, J. W. Olin, E. T. Fung, et al. {beta}2-Microglobulin as a Biomarker in Peripheral Arterial Disease: Proteomic Profiling and Clinical Studies Circulation, September 18, 2007; 116(12): 1396 - 1403. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
G. A. Muller, C. A. Muller, and H. Dihazi Clinical proteomics--on the long way from bench to bedside? Nephrol. Dial. Transplant., May 1, 2007; 22(5): 1297 - 1300. [Full Text] [PDF]
|
|
|
|
 |
|
 D. Fliser, J. Novak, V. Thongboonkerd, A. Argiles, V. Jankowski, M. A. Girolami, J. Jankowski, and H. Mischak Advances in Urinary Proteome Analysis and Biomarker Discovery J. Am. Soc. Nephrol., April 1, 2007; 18(4): 1057 - 1071. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 K. Mosley, F. W. K. Tam, R. J. Edwards, J. Crozier, C. D. Pusey, and L. Lightstone Urinary proteomic profiles distinguish between active and inactive lupus nephritis Rheumatology, December 1, 2006; 45(12): 1497 - 1504. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 T. Pisitkun, R. Johnstone, and M. A. Knepper Discovery of Urinary Biomarkers Mol. Cell. Proteomics, October 1, 2006; 5(10): 1760 - 1771. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
O. H. Ryu, J. C. Atkinson, G. T. Hoehn, G. G. Illei, and T. C. Hart Identification of parotid salivary biomarkers in Sjogren's syndrome by surface-enhanced laser desorption/ionization time-of-flight mass spectrometry and two-dimensional difference gel electrophoresis Rheumatology, September 1, 2006; 45(9): 1077 - 1086. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 G. L. Hortin and B. Meilinger Cross-Reactivity of Amino Acids and Other Compounds in the Biuret Reaction: Interference with Urinary Peptide Measurements Clin. Chem., August 1, 2005; 51(8): 1411 - 1419. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
G. L. Hortin, B. Meilinger, and S. K. Drake Size-Selective Extraction of Peptides from Urine for Mass Spectrometric Analysis Clin. Chem., June 1, 2004; 50(6): 1092 - 1095. [Full Text] [PDF]
|
|
|
|
 |
|
 M. A. Rogers, P. Clarke, J. Noble, N. P. Munro, A. Paul, P. J. Selby, and R. E. Banks Proteomic Profiling of Urinary Proteins in Renal Cancer by Surface Enhanced Laser Desorption Ionization and Neural-Network Analysis: Identification of Key Issues Affecting Potential Clinical Utility Cancer Res., October 15, 2003; 63(20): 6971 - 6983. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Kretzler, C. D. Cohen, P. Doran, A. Henger, S. Madden, E. F. Grone, P. J. Nelson, D. Schlondorff, and H.-J. Grone Repuncturing the Renal Biopsy: Strategies for Molecular Diagnosis in Nephrology J. Am. Soc. Nephrol., July 1, 2002; 13(7): 1961 - 1972. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. A. Knepper Proteomics and the Kidney J. Am. Soc. Nephrol., May 1, 2002; 13(5): 1398 - 1408. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
|
HOME
CURRENT ISSUE
ARCHIVES
JASN Express
ONLINE SUBMISSION
AUTHOR INFO
EDITORIAL BOARD SUBSCRIBE FEEDBACK ALERTS HELP |
