Preparation of peptide-labeled wheat germ agglutinin intended for multiple tracing in a single animal:
Gespeichert in:
| 1. Verfasser: | |
|---|---|
| Format: | Buch |
| Sprache: | German |
| Veröffentlicht: |
1994
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| Schlagworte: | |
| Online-Zugang: | Inhaltsverzeichnis |
| Beschreibung: | Bochum, Univ., Diss., 1995 |
| Beschreibung: | 73 Bl. graph. Darst. |
Internformat
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Datensatz im Suchindex
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CONTENTS
PP
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4
-
1
ABBREVIATIONS
2
INTRODUCTION
2.1 MULTIPLE TRACING TECHNIQUES HAVE BEEN USED FOR
THE VISUALIZATION OF NEURONAL PATHWAYS.
2.2 VARIOUS LECTINS MAY BE SUITED FOR MULTIPLE
TRACING OF PARALLEL PATHWAYS.
2.3 LECTIN-HAPTEN CONJUGATES MAY SHOW VARIOUS
PROBLEMS, WHEN INTENDED FOR MULTIPLE TRACING.
2.4 SMALL PEPTIDES MAY BE INTERESTING CANDIDATES
FOR HAPTENYLATION OF LECTIN TRACERS.
3
MATERIALS AND METHODS
3
#1
PREPARATION OF PEPTIDE-LABELED WGA
3.1.1 CONJUGATION OF PEPTIDES TO WGA
3.1.1.1 ADDITION OF SULFHYDRYL GROUPS TO PEPTIDES
3.1.1.2
DETERMINATION OF SULFHYDRYL GROUP CONTENT
3
.1.1.3 INTRODUCTION OF MALEIMIDO GROUPS INTO WGA
3.1.1.4
DETERMINATION OF INCORPORATED MALEIMIDO GROUPS
AFTER
REACTION WITH SUCCINIMIDYL 6-MALEIMIDO-
HEXANOATE
(MHS)
15
15
15
15
15
16
16
BIBLIOGRAFISCHE INFORMATIONEN
HTTP://D-NB.INFO/944671365
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5
-
3.1.1.5 DETERMINATION OF INCORPORATED MALEIMIDO GROUPS
DURING THE REACTION WITH MALSAC-HNSA
3.1.1.6 CONJUGATION OF SH-PEPTIDES TO MALEIMIDO-WGA 17
3.1.1.7 SEPARATION OF PEPTIDE-LABELED WGA 19
3.1.1.8 DETERMINATION OF INCORPORATED SH-PEPTIDES 19
3.1.1.9 DETERMINATION OF PROTEIN-CONCENTRATIONS 19
3.1.2 ELISA-PROCEDURE FOR THE DETERMINATION OF 20
WGA-ACTIVITY
3.1.3 AFFINITY-PURIFICATION OF PEPTIDE-LABELED WGA 21
3.1.3.1 PREPARATION OF TRISACRYL-OVOMUCOID BEADS 21
3.1.3.2 ADSORPTION AND DESORPTION OF WGA 22
3.2 PREPARATION OF ANTI-PEPTIDE ANTIBODIES 22
FOR DETECTION OF PEPTIDE-LABELED WGA
3.2.1 PREPARATION OF CARRIER-PEPTIDE CONJUGATES OF 22
TWO CONJUGATION-TYPES
3.2.1.1 CONJUGATION WITH GLUTARALDEHYDE 22
3.2.1.2 CONJUGATION WITH IODOACETATE 23
SEPARATION OF IODOACETYLATED BSA FROM EXCESS
IODOACETIC ACID
DETERMINATION OF INCORPORATED IODOACYL GROUPS
COUPLING OF IMINOTHIOLANE-MODIFIED PEPTIDES TO
IODOACETYLATED BSA
3.2.2 PREPARATION OF ANTI-PEPTIDE-ANTIBODIES 26
3.2.2.1 IMMUNIZATION WITH CARRIER-PEPTIDE-CONJUGATES OF 26
TWO LINKAGE TYPES AND RAISING OF ANTIBODIES
3.2.2.2 AFFINITY PURIFICATION AND CHARACTERIZATION OF 26
ANTISERA
3.2.2.3 AFFINITY BIOTINYLATION OF THE
28
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6
-
ANTI-GA-PEPTIDE-ANTIBODIES
3.2.2.4 PREPARATION OF IGG-FRACTIONS 28
3.2.3 STAINING OF PEPTIDES IN ARTIFICIAL TISSUES 29
4
RESULTS
30
4.1 CONJUGATION OF PEPTIDES TO WGA
31
4.1.1 ADDITION OF SH-GROUPS TO THE PEPTIDES 33
4.1.2 INTRODUCTION OF MALEIMIDO GROUPS INTO WGA 35
PROVED TO BE MOST SUCCESSFUL WITH THE N-HYDROXY-
SUCCINIMIDE ESTER OF MALEIMIDOHEXANOIC ACID (MHS)
4.1.3 CONJUGATION OF SH-PEPTIDES TO MALEIMIDO-WGA 40
4.2 BLISA-ASSAY FOR DETERMINATION OF 42
WGA-ACTIVITY
4.2.1 DEVELOPMENT OF THE ELISA WITH UNCONJUGATED WGA 42
4.2.2 PEPTIDE-LABELED WGA WAS DETECTED WITH 45
ANTI-PEPTIDE-ANTIBODIES
4.2.3 PARAMETERS FOR THE PEPTIDE-CONJUGATION OF WGA 48
RESULTING IN OPTIMAL ANTI-PEPTIDE-SIGNALS WITH
MINIMAL LOSS OF LECTIN ACTIVITY WERE DETERMINED
WITH THE NEWLY ELISA
4.3 AFFINITY PURIFICATION AND PROPERTIES OF
51
PEPTIDE-WGA
4.3.1 PEPTIDE-MOLECULES WITH RETAINED LECTIN ACTIVITY 51
WERE SEPARATED FROM INACTIVATED MOLECULES BY
AFFINITY PURIFICATION
4.3.2 AFFINITY PURIFIED GGY-WGA SHOWED THE HIGHEST 53
ELISA-SENSITIVITY CLOSELY FOLLOWING THE UNLABELED
WGA
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7
-
5
DISCUSSION 57
5.1 RATIONAL FOR THE PREPARATION
OF PEPTIDE-LABELED WGA. 57
5.2 NEW, HIGHLY SENSITIVE ELISA FOR THE
DETERMINATION OF LECTIN ACTIVITY OF
NATIVE AND CONJUGATED WGA. 57
5.3 CONJUGATION OF GGY TO WGA PROVED MOST
SUCCESSFUL WITH THE N-HYDROXYSUCCINIMIDE
ESTER OF MALEIMIDO HEXANOIC ACID. 59
5.4
SPECIFIC ANTI-PEPTIDE ANTIBODIES ALLOW
INDEPENDENT VISUALIZATION OF THE GGY- AND
GGR-LABELED WGA TRACER MOLECULES. 61
5.5
PROPERTIES AND APPLICABILITY OF THE NEW
PEPTIDE-LABELED WGA. 62
T
6
SUMMARY 63
7
REFERENCES 64 |
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| building | Verbundindex |
| bvnumber | BV010334241 |
| ctrlnum | (OCoLC)64525682 (DE-599)BVBBV010334241 |
| discipline | Medizin |
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| illustrated | Illustrated |
| indexdate | 2024-10-09T16:17:25Z |
| institution | BVB |
| language | German |
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| physical | 73 Bl. graph. Darst. |
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| spelling | Czekalla, Jörg Verfasser aut Preparation of peptide-labeled wheat germ agglutinin intended for multiple tracing in a single animal vorgelegt von Jörg Czekalla 1994 73 Bl. graph. Darst. txt rdacontent n rdamedia nc rdacarrier Bochum, Univ., Diss., 1995 (DE-588)4113937-9 Hochschulschrift gnd-content DNB Datenaustausch application/pdf http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=006877650&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA Inhaltsverzeichnis |
| spellingShingle | Czekalla, Jörg Preparation of peptide-labeled wheat germ agglutinin intended for multiple tracing in a single animal |
| subject_GND | (DE-588)4113937-9 |
| title | Preparation of peptide-labeled wheat germ agglutinin intended for multiple tracing in a single animal |
| title_auth | Preparation of peptide-labeled wheat germ agglutinin intended for multiple tracing in a single animal |
| title_exact_search | Preparation of peptide-labeled wheat germ agglutinin intended for multiple tracing in a single animal |
| title_full | Preparation of peptide-labeled wheat germ agglutinin intended for multiple tracing in a single animal vorgelegt von Jörg Czekalla |
| title_fullStr | Preparation of peptide-labeled wheat germ agglutinin intended for multiple tracing in a single animal vorgelegt von Jörg Czekalla |
| title_full_unstemmed | Preparation of peptide-labeled wheat germ agglutinin intended for multiple tracing in a single animal vorgelegt von Jörg Czekalla |
| title_short | Preparation of peptide-labeled wheat germ agglutinin intended for multiple tracing in a single animal |
| title_sort | preparation of peptide labeled wheat germ agglutinin intended for multiple tracing in a single animal |
| topic_facet | Hochschulschrift |
| url | http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=006877650&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA |
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