Mouse anti-Actin (Smooth muscle), Clone 1A4
Contents
1. REQUISITOS PARA LA TINCI N INMUNOHISTOQU MICA El prediluido V Line de Invitrogen fue creado para usarse con los sistemas de tinci n inmunohistoqu mica Ventana EST NexES y BenchMark juntamente con los Equipos de detecci n Ventana y los dispensadores de reactivo Ventana Invitrogen ha realizado la valoraci n y controlado la calidad de este anticuerpo para garantizar un rendimiento uniforme y fiable La Tabla 1 resume los requisitos para la inmunotinci n Tabla 1 Requisitos para Tejido FFPE Especificaci n Eliminaci n de Ep tope Inducida por Calor HIER No Necesaria Digesti n enzim tica No Necesaria Incubaci n enzim tica No Necesaria Incubaci n del anticuerpo primario 32 minutos www invitrogen com Invitrogen Corporation 542 Flynn Rd Camarillo CA 93012 Tel 800 955 6288 E mail techsupport invitrogen com PI 08 4106 Rev 08 08 DCC 08 1089 2007 Invitrogen Corporation All right reserved These products may be covered by one or more Limited Use Label Licenses see the Invitrogen Catalog or www invitrogen com By use of these products you accept the terms and conditions of all applicable Limited Use Label Licenses MATERIALES NECESARIOS PERO NO SUMINISTRADO Reactivo N de catalogo 1 HistoGrip M 00 8050 2 PBS PBS 0 01 M 00 3000 3 Hematoxilina de Mayer 00 8011 4 Tamp n citrato pH 6 0 si se necesita para HIER 00 5000 5 Soluci n EDTA si se necesita para HIE2. EC Invitrogen Ltd Inchinnan Business Park 3 Fountain Drive Paisley PA4 9RF UK www invitrogen com Invitrogen Corporation 542 Flynn Rd Camarillo CA 93012 Tel 800 955 6288 E mail techsupport invitrogen com PI 08 4106 Rev 08 08 DCC 08 1089 2007 Invitrogen Corporation All right reserved These products may be covered by one or more Limited Use Label Licenses see the Invitrogen Catalog or www invitrogen com By use of these products you accept the terms and conditions of all applicable Limited Use Label Licenses e invitrogen CE Anticuerpo Actin de rat n Clon 1 A4 Para utilizaci n en diagn stico in vitro X 08 4106 10 mL Anticuerpo prediluido V Line listo para usar Optimizado para los sistemas de tinci n inmunohistoqu mica Ventana ES NexES y BenchMark PROP SITO DE USO Para utilizaci n en diagn stico in vitro La interpretaci n de los resultados debe realizarla un pat logo cualificado dentro del contexto de la historia cl nica del paciente y otras pruebas diagn sticas REACTIVO SUMINISTRADO El anticuerpo Actin de rat n se purifica a partir de fluido de ascitis en rat n y se diluye en tamp n fosfato PBS Ph 7 4 y seroalb mina bovina al 1 BSA con azida de sodio al 0 1 NaN como conservante Inmunog n P ptido de la actina sint tica Isotipo IgG Clon 1A4 Control positivo Leiomiosarcoma Pauta de tinci n esperada Citoplasm tico ALMACENAMIENTO 2 8 C
3. H matoxylin 1 2 Minuten Mehrmals in Leitungswasser waschen bis das Wasserbad sauber bleibt Zweimal in Blaul sung platzieren z B 10 mM PBS pH 7 4 15 20 Sekunden Zweimal in destilliertem Wasser waschen Insgesamt 3 Minuten Nacheinander in 70 95 95 100 und 100 EtOH dehydrieren Dann in Xylen und Xylen dehydrieren Bei Verwendung von AEC nicht dehydrieren Je 2 Minuten Deckglas mit Histomount Fixativ anbringen f r AEC muss GVA Fixativ verwendet werden Vor der Betrachtung trocknen lassen 10 Minuten FEHLERBEHEBUNG Problem L sung Positive Kontrolle weist eine schwache Immunf rbung auf i Reagenzien ist Andere positive Kontrollen die w hrend desselben Verfahrens pr pariert wurden pr fen um zu bestimmen ob die Ursache der prim re Antik rper oder eines der allgemeinen sekund ren Positive Kontrolle ist negativ eines der allgemeinen sekund ren Reagenzien ist Sicherstellen dass der Objekttr ger mit dem richtigen Ventana Strichcode Etikett gekennzeichnet ist Bei richtiger Etikettierung m ssen andere positive Kontrollen die w hrend desselben Verfahrens pr pariert wurden gepr ft werden um zu bestimmen ob die Ursache der prim re Antik rper oder WARENZEICHEN Cap Plus Clearmount Digest AIITM HistoGrip Histomount und Histostain NBA PicTure und Zymed sind Warenzeichen der Zymed Laboratories Inc
4. antes de la desparafinaci n de los portaobjetos puesto que pueden pasar varios minutos antes de que hierva el tamp n 4 Colocar los portaobjetos en una rejilla de soporte para portaobjetos y ponerlos en el vaso de precipitados con el tamp n hirviendo D jelo hervir durante 15 minutos 5 Despu s de calentar retire el vaso de precipitado de la placa de calor y d jelo que se enfr e a temperatura ambiente durante al menos 15 20 minutos 6 Enjuagar los portaobjetos con PBS n de cat 00 3000 y proceda con el protocolo de inmunotinci n Digesti n enzim tica en caso necesario 1 Consulte la Tabla 1 para ver el requisito espec fico de digesti n enzim tica y el tiempo de incubaci n CARGA DEL DISPENSADOR DEFINIDO POR EL USUARIO Invitrogen recomienda el siguiente procedimiento para llenar los dispensadores de reactivo Ventana con el prediluido V Line de Invitrogen Consulte el Manual del Usuario de Ventana para obtener m s informaci n 1 Retire el tap n del extremo de la Jeringa de transferencia Ventana luego retire el mbolo por completo del cilindro y vuelva a colocar el tap n en el cilindro de la jeringa 2 Coloque una etiqueta a la Jeringa de reactivo y transferencia Ventana indicando el anticuerpo Invitrogen utilizado Vierta la ampolla que se adjunta de prediluido V Line 10 mL anticuerpo listo para usar en el cilindro de la Jeringa de transferencia Ventana luego vuelva a colocar el mbolo en el cil
5. di trasferimento Ventana specificando il nome dell anticorpo Invitrogen utilizzato 3 Versare il contenuto della boccetta acclusa di anticorpo prediluito V Line di Invitrogen 10 mL anticorpo pronto per l uso nel cilindro della siringa di trasferimento Ventana quindi reinserire lo stantuffo nel cilindro 4 Capovolgere la siringa di trasferimento Ventana in modo che il tappo sia rivolto verso l alto e lo stantuffo verso il basso Quindi rimuovere il tappo Eliminare l aria dalla siringa di trasferimento Ventana spingendo cautamente lo stantuffo verso il tappo A questo punto rimuovere il tappo dalla punta della siringa di trasferimento Ventana Collegare l ugello della siringa per reagenti Ventana all ugello della siringa di trasferimento Ventana Erogare l anticorpo dalla siringa di trasferimento Ventana alla siringa per reagenti Ventana La siringa per reagenti Ventana ha una capacit massima di 12 5 mL ovvero un quantitativo sufficiente per eseguire 120 prove circa 9 Attaccare l etichetta acclusa Invitrogen sulla siringa per reagenti Ventana BE mn L anticorpo prediluito V Line di Invitrogen fornito pronto per l uso ed da utilizzarsi con i sistemi di colorazione immunoistochimica Ventana ES NexES e BenchMark Per informazioni particolareggiate sul funzionamento e la sequenza procedurale dei sistemi di colorazione immunoistochimica Ventana EST NexES e BenchMark si pr
6. per immunocolorazione debole stabilire se ci causato dall anticorpo primario o da uno dei reagenti secondari comuni Il controllo positivo negativo reagenti secondari comuni Accertarsi che il vetrino abbia l etichetta con il codice a barre Ventana corretta Qualora sia etichettato correttamente controllare altri controlli positivi preparati durante lo stesso ciclo di azionamento del dispositivo per stabilire se ci causato dall anticorpo primario o da uno dei MARCHI REGISTRATI Cap Plus Clearmount Digest All HistoGrip Histomount e Histostain NBATM PicTure e Zymed sono marchi registrati di Zymed Laboratories Inc Ventana ES NexES e BenchMark sono marchi commerciali di propriet di Ventana Medical Systems Inc Gli anticorpi V Line sono sviluppati esclusivamente da Zymed Laboratories Inc e non implicano alcuna autorizzazione o approvazione da parte di Ventana Medical Systems Inc Zymed e Ventana non sono imprese consociate affiliate o collegate in alcun altro modo Per informazioni pi dettagliate vi preghiamo di consultare la versione inglese del catalogo oppure visitate il sito www invitrogen com SE AVETE DOMANDE SUL PRODOTTO CONTATTATE IL VOSTRO DISTRIBUTORE PI VICINO Rappresentante Autorizzato per IVDD 98 79 EC Invitrogen Ltd Inchinnan Business Park 3 Fountain Drive Paisley PA4 9RF UK www invitrogen com Invitrogen Corporation 54
7. 00 and 100 EtOH Then in xylene and xylene Do not dehydrate 2 Minutes Each when using AEC Apply coverslip with HistomountTM Mounting Solution Use GVA Mounting Solution when using AEC Allow to dry before viewing 10 Minutes TROUBLESHOOTING Problem Solution Positive control shows weak Check other positive controls prepared during the same instrument run to determine if it is due to the immunostaining primary antibody or to one of the common secondary reagents Assure that the slide has proper Ventana Bar Coding Label If it is labeled properly check other Positive control is negative positive controls prepared during the same instrument run to determine if it is due to the primary antibody or one of the common secondary reagents REFERENCES 1 GalR et al Cell Vision 2 2 129 1995 2 Tsukada T et al Amer J Pathology 126 51 127 389 1987 3 Rangdaeng S et al Amer J Clin Pathology 96 1 32 45 1991 TRADEMARKS Cap Plus Clearmount Digest AIITM HistoGrip Histomount and Histostain NBA PicTure and Zymed are trademarks of Zymed Laboratories Inc Ventana ES NexES and BenchMark are trademarks of Ventana Medical Systems Inc V Line antibodies are developed solely by Zymed Laboratories Inc and do not imply approval or endorsement by Ventana Medical Systems Inc Zymed and Ventana are not affiliated associated or related in any way Authorized Representative for VDD 98 79
8. 2 Flynn Rd Camarillo CA 93012 Tel 800 955 6288 E mail techsupport invitrogen com PI 08 4106 Rev 08 08 DCC 08 1089 2007 Invitrogen Corporation All right reserved These products may be covered by one or more Limited Use Label Licenses see the Invitrogen Catalog or www invitrogen com By use of these products you accept the terms and conditions of all applicable Limited Use Label Licenses e invitrogen CE Maus Anti Actin Klon 1A4 F r In Vitro Diagnostischen Gebrauch X 08 4106 10 mL V Line Vorverdiinnung gebrauchsfertiger Antik rper Optimiert f r die immunhistochemischen F rbungssysteme Ventana ES NexES und BenchMark BEABSICHTIGTER NUTZEN Fiir In Vitro Diagnostischem Gebrauch Die Interpretation muss im Zusammenhang mit der klinischen Vorgeschichte des Patienten und anderen Diagnostiktests eines qualifizierten Pathologen vorgenommen werden REAGENS GELIEFERT Maus Anti Actin ist gereinigt von Maus asciter Fl ssigkeit und verd nnt in Phosphatgepufferter Salzl sung PBS pH 7 4 und 1 igem Rinder Serum Albumin BSA mit 0 1 Sodium Azid NaN als ein Konservierungsmittel Immunogen Synthetisches Actin Peptid Isotyp IgG Klon 144 Positive Kontrolle Leiomyosarkom Erwartetes F rbungsmuster Cytoplasmatisch LAGERUNG 2 8 C VORAUSSETZUNGEN F R DIE IMMUNHISTOCHEMISCHE FARBUNG Invitrogens V Line Vorverd nnung wurde speziell f r die Verwendung mit den immunhistochemi
9. ALI NECESSARI MA NON FORNITI Reagente Catalogo n 1 HistoGrip 00 8050 2 PBS 0 01 M PBS 00 3000 3 Ematossilina di Mayer 00 8011 4 Soluzione tamponata a base di citrato pH 6 0 se necessaria per il HIER 00 5000 5 Soluzione a base di EDTA se necessaria per il HIER 00 5500 6 Sistemi di colorazione immunoistochimica Ventana EST NexES e BenchMark e Etichette con codice a barre e Coloratori automatici di vetrini EST NexES o BenchMark e Kitdirilevazione DAB o AEC e Software specifico per il kit di rilevazione e Soluzione di lavaggio e Reagente controllo negativo e Coprivetrino liquido 7 Soluzione montante Histomount per DAB Cat n 00 8030 GVA per AEC oppure rosso solido Cat n 00 8000 oppure Clearmount M per AEC DAB oppure rosso solido Cat n 00 8010 PREPARAZIONE DEI CAMPIONI 1 Utilizzare regolarmente tessuto fissato in 10 formalina neutra tamponata oppure in altro fissativo o in alternativa sezioni di tessuto congelato 2 Tagliare sezioni da 3 4 um e collocarle su vetrini caricati positivamente 3 Lasciare asciugare durante la notte a 37 C oppure per 2 4 ore a 58 C PRETRATTAMENTO Heat Induced Epitope Retrieval HIER Recupero di epitopi mediante calore se richiesto 1 Deparaffinizzare i vetrini Le sezioni dei tessuti dovrebbero essere montate su vetrini rivestiti di silano poly L Lysine oppure HistoGrip Cat n 00 8050 2 Lavare i vetrini con acqua distillata per 3 pe
10. EFINI PAR L UTILISATEUR Invitrogen recommande l utilisation de la proc dure ci apr s pour remplir les distributeurs de r actif Ventana avec la pr dilution V Line de Invitrogen Se reporter au manuel de l utilisateur de Ventana pour des d tails suppl mentaires 1 Retirer le bouchon l extr mit de la seringue de transfert Ventana retirer ensuite compl tement le piston du corps de pompe et remettre le bouchon dans le corps de pompe de la seringue 2 Marquer de fa on appropri le r actif Ventana et la seringue de transfert avec l anticorps de Invitrogen utilis 3 Verser le flacon de pr dilution V Line inclus 10 mL anticorps pr t l emploi dans le corps de pompe de la seringue de transfert de Ventana ensuite remettre le piston dans le corps de pompe 4 Retourner la seringue de transfert de Ventana le bouchon tourn vers le haut et le piston vers le bas Retirer ensuite le bouchon Chasser l air de la seringue de transfert Ventana en poussant doucement le piston vers le bouchon Retirer ensuite le bouchon de l extr mit de la seringue de r actif Ventana Attacher l embout de la seringue de r actif Ventana sur l embout de la seringue de transfert Ventana Faire passer l anticorps de la seringue de transfert Ventana dans la seringue de r actif Ventana La quantit maximale contenue dans la seringue de r actif Ventana est de 12 5 mL ce qui permet d effectuer environ 120 t
11. IER Pas Requis Enzyme de digestion Pas Requis Enzyme d Incubation Pas Requis Incubation des anticorps primaires 32 minutes www invitrogen com Invitrogen Corporation 542 Flynn Rd Camarillo CA 93012 Tel 800 955 6288 E mail techsupport invitrogen com PI 08 4106 Rev 08 08 DCC 08 1089 2007 Invitrogen Corporation All right reserved These products may be covered by one or more Limited Use Label Licenses see the Invitrogen Catalog or www invitrogen com By use of these products you accept the terms and conditions of all applicable Limited Use Label Licenses MATERIELS NECESSAIRES MAIS PAS FOURNIS R actif Catalogue No 1 HistoGrip 00 8050 2 PBS 0 01 M PBS 00 3000 3 Mayer H matoxyline 00 8011 4 Tampon citrate pH 6 0 si la RAC l exige 00 5000 5 Solution EDTA si la RAC l exige 00 5500 6 Syst mes de coloration immunohistochimique Ventana EST NexES ou BenchMark e Etiquettes code barres e Marqueur de lames automatis ESM NexES ou BenchMark e Kit de d tection DAB ou AEC e Logiciel sp cifique au kit de d tection e Solution de lavage e R actif de t moin n gatif e Couvre lames liquide 7 Solution de Monture Histomount pour DAB Cat No 00 8030 GVA pour AEC ou Rouge Fixe Cat No 00 8000 ou Clearmount pour AEC DAB ou Rouge Fixe Cat No 00 8010 PREPARATION DE SPECIMENS 1 Utiliser un tissu fix dans 10 Formaline Neutre Tamponn e
12. Pour des informations plus d taill es veuillez vous r f rer aux feuilles de donn es en version anglaise ou aller sur www invitrogen com SI VOUS AVEZ DES QUESTIONS SUR CE PRODUIT CONTACTEZ VOTRE DISTRIBUTEUR LOCAL Repr sentant Autoris pour IVDD 98 79 EC Invitrogen Ltd Inchinnan Business Park 3 Fountain Drive Paisley PA4 9RF UK www invitrogen com Invitrogen Corporation 542 Flynn Rd Camarillo CA 93012 Tel 800 955 6288 E mail techsupport invitrogen com PI 08 4106 Rev 08 08 DCC 08 1089 2007 Invitrogen Corporation All right reserved These products may be covered by one or more Limited Use Label Licenses see the Invitrogen Catalog or www invitrogen com By use of these products you accept the terms and conditions of all applicable Limited Use Label Licenses e invitrogen CE Topo anti Actin Clone 1A4 Per uso diagnostico In Vitro X 08 4106 10 mL Anticorpo prediluito V Line Pronto per l uso Ottimizzato per i sistemi di colorazione immunoistochimica Ventana ES NexES e BenchMark SCOPI D UTILIZZO Per uso diagnostico In Vitro L interpretazione deve essere effettuata da un patologo qualificato entro il contesto dell anamnesi clinica del paziente ed in considerazione di altri test diagnostici REAGENTI FORNITI Topo anti Actin viene purificato da liquido ascitico di topo e diluito in soluzione salina tamponata al fosfato PBS pH 7 4 e 1 di albumina di siero bovino BSA co
13. R 00 5500 6 Sistemas de tinci n inmunohistoqu mica Ventana EST NexES o BenchMark e Etiquetas de c digos de barras e Tinte de portaobjetos automatizado EST NexES o BenchMark e Equipo de detecci n DAB o AEC e Software espec fico para el Equipo de detecci n e Soluci n de lavado e Reactivo para control negativo e Cubreobjetos l quido 7 Soluci n de montaje Histomount para DAB n de cat 00 8030 GVA para AEC o para Fast Red n de cat 00 8000 o Clearmount M para AEC DAB o Fast Red n de cat 00 8010 PREPARACI N DE LA MUESTRA 1 Utilizar tejido fijado en formalina neutra tamponada al 10 u otro fijador general o cortes de tejido congelados 2 Hacer cortes de 3 4 um y colocarlos sobre los portaobjetos cargados positivamente 3 Secar durante toda la noche a 37 C o de 2 a 4 horas a 58 C PRETRATAMIENTO Eliminaci n de Ep tope Inducida por Calor HIER en caso necesario 1 Proceder con la desparafinaci n de las muestras Los cortes de tejido deber an montarse sobre portaobjetos cubiertos de silano poli L lisina o HistoGrip n de cat 00 8050 2 Lavar los portaobjetos con agua destilada 3 veces durante 2 minutos cada vez 3 Poner un vaso de precipitados Pyrex de 1 litro con 500 mL de tamp n de citrato 0 01 M n de cat 00 5000 o soluci n de EDTA n de cat 00 5500 en una placa de calor Calentar la soluci n tamp n hasta que hierva Este paso se puede preparar
14. S 0 01 M PBS 00 3000 4 Citrate Buffer pH 6 0 if required for HIER 00 5000 5 EDTA Solution if required for HIER 00 5500 6 Ventana ESTM NexES or BenchMark Immunohistochemistry Staining Systems e Bar Code Labels e ESTM NexES or BenchMark Automated Slide Stainer e DAB or AEC Detection Kit e Detection Kit specific software e Wash Solution e Negative Control Reagent e Liquid Coverslip 7 Mounting solution Histomount for DAB Cat No 00 8030 GVA for AEC or Fast Red Cat No 00 8000 or Clearmount for AEC DAB or Fast Red Cat No 00 8010 SPECIMEN PREPARATION 1 Use tissue fixed in 10 neutral buffered formalin or other fixative on regular basis or frozen tissue sections 2 Cut 3 4 um sections and place on positively charged slides 3 Dry overnight at 37 C or for 2 4 hrs at 58 C PRETREATMENT Heat Induced Epitope Retrieval HIER if required 1 Deparaffinize slides Tissue sections should be mounted on silane poly L Lysine or HistoGrip Cat No 00 8050 coated slides 2 Wash slides with distilled water 3 times for 2 min each 3 Place a IL glass Pyrex beaker containing 500 mL of 0 01 M citrate buffer Cat No 00 5000 or EDTA solution Cat No 00 5500 on a hot plate Heat the buffer solution until it boils This step may be prepared before slide deparaffinization as the buffer may take several minutes to boil 4 Put the slides in a slide rack and place in the beaker with boili
15. Ventana ES NexES und BenchMark sind Marken der Ventana Medical Systems Inc V Line Antik rper werden ausschlieBlich von Zymed Laboratories Inc entwickelt und wurden nicht durch Ventana Medical Systems Inc genehmigt oder gef rdert Zymed und Ventana sind in keiner Weise angegliedert aneinander angeschlossen bzw miteinander verbunden F r genauere Information beziehen Sie sich bitte entweder auf die englische Version auf der Datenplatte oder auf www invitrogen com SOLLTEN SIE IRGENDWELCHE FRAGEN BER DIESES PRODUKT HABEN WENDEN SIE SICH BITTE AN IHREN RTLICHEN VERTEILER Bevollm chtigter Repr sentant f r IVDD 98 79 EC Invitrogen Ltd Inchinnan Business Park 3 Fountain Drive Paisley PA4 9RF UK www invitrogen com Invitrogen Corporation 542 Flynn Rd Camarillo CA 93012 Tel 800 955 6288 E mail techsupport invitrogen com PI 08 4106 Rev 08 08 DCC 08 1089 2007 Invitrogen Corporation All right reserved These products may be covered by one or more Limited Use Label Licenses see the Invitrogen Catalog or www invitrogen com By use of these products you accept the terms and conditions of all applicable Limited Use Label Licenses
16. e Invitrogen Catalog or www invitrogen com By use of these products you accept the terms and conditions of all applicable Limited Use Label Licenses CONTRATINCI N Y CONSERVACI N Protocolo Tiempo Contratinci n con hematoxilina 1 2 minutos Lavar en agua corriente con varios cambios hasta que el ba o de agua permanezca limpio Colocar en soluci n azulante es decir 10 mM PBS pH 7 4 con 2 cambios 15 20 segundos Lavar en agua destilada con 2 cambios 3 minutos en total Deshidratar secuencialmente en EtOH 70 95 95 100 y 100 Luego en xileno y xileno no deshidratar al usar AEC 2 minutos cada uno Aplicar cubreobjetos con soluci n de montaje Histomount Usar soluci n de montaje GVA cuando se use AEC Dejar secar antes de ver 10 minutos DETECCI N Y DIAGN STICO DE AVER AS Problema Soluci n El control positivo muestra una Verifique los dem s controles positivos preparados durante esa misma serie con el instrumento para inmunotinci n d bil determinar si se debe al anticuerpo primario o a uno de los reactivos secundarios comunes Aseg rese de que el portaobjetos tenga la etiqueta con el c digo de barras de Ventana correcta Si est etiquetado correctamente verifique los dem s controles positivos preparados durante esa misma serie con el instrumento para determinar si se debe al anticuerpo primario o a uno de los reactivos secundarios comunes El control pos
17. e invitrogen CE Mouse anti Actin Smooth muscle Clone 1 A4 For In Vitro Diagnostic Use Lot No X 08 4106 10 mL V Line Predilute Ready to Use Antibody Optimized For Ventana ES NexES and BenchMark Immunohistochemistry Staining Systems INTENDED USE For In Vitro Diagnostic Use Invitrogen s Mouse anti Actin Smooth muscle antibody is intended to qualitatively stain actin protein in frozen and formalin fixed paraffin embedded tissue sections Interpretation must be made within the context of the patient s clinical history and other diagnostic tests by a qualified pathologist BACKGROUND Actin is a highly conserved skeletal protein which is in both muscle and nonmuscle cells The mammalian tissues express at least 6 unique actin isotypes Alpha isotype is found exclusively in muscle cells including alpha skeletal alpha cardiac and alpha and gamma smooth muscle actin Beta and gamma nonmuscular actins have been observed in nonmuscle cells This antibody is specific for the alpha isoform of smooth muscle actin 1A4 does not cross react with skeletal or cardiac muscle actin mesenchymal cells epithelial cells or other cytoskeletal components This antibody reacts with smooth muscle actin REAGENT PROVIDED Mouse anti Actin Antibody is purified from mouse ascites and diluted in phosphate buffered saline PBS pH 7 4 and 1 bovine serum albumin BSA with 0 1 sodium azide NaN as a preservative Immunog
18. ega di consultare i rispettivi Manuali per l utente Ventana www invitrogen com Invitrogen Corporation 542 Flynn Rd Camarillo CA 93012 Tel 800 955 6288 E mail techsupport invitrogen com PI 08 4106 Rev 08 08 DCC 08 1089 2007 Invitrogen Corporation All right reserved These products may be covered by one or more Limited Use Label Licenses see the Invitrogen Catalog or www invitrogen com By use of these products you accept the terms and conditions of all applicable Limited Use Label Licenses CONTROCOLORAZIONE E CONSERVAZIONE Protocollo Tempo Controcolorare con Ematossilina 1 2 minuti Lavare in acqua di rubinetto ricambiata diverse volte finch il bagnomaria rimane limpido Immergere in soluzione azzurrante ovvero 10 mM di PBS pH 7 4 eseguendo 2 ricambi 15 20 secondi Lavare in acqua distillata eseguendo 2 ricambi 3 minuti in tutto Disidratare sequenzialmente in EtOH al 70 95 95 100 e 100 Quindi in xilene e xilene Non disidratare se si usa 1 AEC 2 minuti ciascuno Applicare il coprivetrino con la Soluzione di fissaggio Histomount Se si usa I AEC utilizzare la Soluzione di fissaggio GVA Lasciare asciugare prima dell osservazione 10 minuti RISOLUZIONE DEI PROBLEMI Problema Soluzione Il controllo positivo mostra una Verificare altri controlli positivi preparati durante lo stesso ciclo di azionamento del dispositivo
19. en Actin synthetic peptide Total protein concentration g L Clone 1A4 Isotype IgG Antibody concentration mg L STORAGE 2 8 C POSITIVE CONTROL TISSUE Leiomyosarcoma EXPECTED STAINING PATTERN Cytoplasmic REQUIREMENTS FOR IMMUNOHISTOCHEMISTRY STAINING Invitrogen s V Line Predilute was developed for use with the Ventana ESTM NexES and BenchMarkTM Immunohistochemistry Staining Systems in combination with Ventana Detection Kits and Ventana Reagent Dispensers Invitrogen has titered and quality controlled this antibody to assure consistent and reliable performance Table 1 summarizes the requirements for immunostaining Table 1 Requirement for FFPE Tissues Specification HIER Heat Induced Epitope Retrieval Not Required Enzyme Digestion Not Required Enzyme Incubation Not Required Primary Antibody Incubation 32 minutes www invitrogen com Invitrogen Corporation 542 Flynn Rd Camarillo CA 93012 Tel 800 955 6288 E mail techsupport invitrogen com PI 08 4106 Rev 08 08 DCC 08 1089 2007 Invitrogen Corporation All right reserved These products may be covered by one or more Limited Use Label Licenses see the Invitrogen Catalog or www invitrogen com By use of these products you accept the terms and conditions of all applicable Limited Use Label Licenses MATERIALS REQUIRED BUT NOT PROVIDED Reagent Catalog No 1 HistoGrip 00 8050 2 Antibody Diluent 00 3118 3 PB
20. en Verschluss entfernen Zum Entl ften der Ventana Transferspritze den Kolben vorsichtig in Richtung Verschluss schieben Dann den Verschluss von der Spitze der Ventana Reagenzspritze abnehmen Die Ventana Reagenzspritzend se an der Ventana Transferspritzend se befestigen Antik rper aus der Ventana Transferspritze in die Ventana Reagenzspritze spritzen Das maximale Fassungsverm gen der Ventana Reagenzspritze betr gt 12 5 mL und reicht f r etwa 120 Tests Das beiliegende Invitrogen Etikett auf der Ventana Reagenzspritze anbringen www invitrogen com Invitrogen Corporation 542 Flynn Rd Camarillo CA 93012 Tel 800 955 6288 E mail techsupport invitrogen com Rev 08 08 DCC 08 1089 2007 Invitrogen Corporation All right reserved These products may be covered by one or more Limited Use Label Licenses see the Invitrogen Catalog or www invitrogen com By use of these products you accept the terms and conditions of all applicable Limited Use Label Licenses Der Invitrogen V Line Vorverd nnungs Antik rper kann jetzt mit den immunhistochemischen F rbungssystemen Ventana ES NexES und BenchMark eingesetzt werden Spezifische Details zum Betrieb und zur Ereignissequenz der immunhistochemischen F rbungssysteme Ventana ES NexES und BenchMark sind den entsprechenden Ventana Benutzerhandbiichern zu entnehmen GEGENFARBUNG UND KONSERVIERUNG Protokoll Zeit Gegenf rbung mit
21. ests 9 Poser l tiquette de Invitrogen ci jointe sur la seringue de r actif de Ventana DO ONO L anticorps de pr dilution V Line de Invitrogen est pr t tre utilis avec les syst mes de coloration immunohistochimique de Ventana ES NexES et BenchMark Pour des d tails sp cifiques concernant le fonctionnement et la s quence des v nements des syst mes de coloration immunohistochimique de Ventana EST NexES et BenchMark se reporter au manuel de l utilisateur Ventana en question www invitrogen com Invitrogen Corporation 542 Flynn Rd Camarillo CA 93012 Tel 800 955 6288 E mail techsupport invitrogen com PI 08 4106 Rev 08 08 DCC 08 1089 2007 Invitrogen Corporation All right reserved These products may be covered by one or more Limited Use Label Licenses see the Invitrogen Catalog or www invitrogen com By use of these products you accept the terms and conditions of all applicable Limited Use Label Licenses CONTRECOLORATION ET CONSERVATION Protocole Dur e Contremarquer l h matoxyline 1 2 minutes Laver plusieurs reprises avec de l eau du robinet jusqu ce l eau soit bien claire Placer dans une solution d azurage par exemple PBS 10 mM pH 7 4 deux reprises 15 20 secondes Laver dans de l eau distill e deux reprises 3 minutes en tout D shydrater ensuite dans de l thanol 70 95 95 100 e
22. ge into the Ventana Reagent Syringe The maximum amount held by the Ventana Reagent Syringe is 12 5 mL which provides approximately 120 tests 9 Place the enclosed Invitrogen label onto the Ventana Reagent Syringe CON EE Invitrogen s V Line Predilute antibody is ready for use with the Ventana ES NexES and BenchMark Immunohistochemistry Staining Systems For specific details on operation and sequence of events of the Ventana ES NexES and BenchMark Immunohistochemistry Staining Systems refer to the respective and appropriate Ventana Operator s Manual www invitrogen com Invitrogen Corporation 542 Flynn Rd Camarillo CA 93012 Tel 800 955 6288 E mail techsupport invitrogen com PI 08 4106 Rev 08 08 DCC 08 1089 2007 Invitrogen Corporation All right reserved These products may be covered by one or more Limited Use Label Licenses see the Invitrogen Catalog or www invitrogen com By use of these products you accept the terms and conditions of all applicable Limited Use Label Licenses COUNTERSTAINING amp PRESERVATION Protocol Time Counterstain with hematoxylin 1 2 Minutes Wash in tap water with several changes until water bath remains clear Place in blueing solution i e 10 mM PBS pH 7 4 with 2 changes 15 20 Seconds Wash in distilled water with 2 changes 3 Minutes Total pu 1 i Le oj Le i pu Dehydrate sequentially in 70 95 95 1
23. iedergewinnung der Epitope erforderlich 00 5500 6 Immunhistochemische F rbungssysteme Ventana EST NexES oder BenchMark e Strichcode Etikett e Objekttriger F rbeautomaten ES NexES oder BenchMark e DAB oder AEC Detektionskit e Detektionskit spezifische Software e Waschl sung e Negatives Kontrollreagenz e Fliissigkeits Deckglas 7 Pr parierungsl sung Histomount f r DAB Cat No 00 8030 GVA f r AEC oder Lichtstark Rot Cat Nr 00 8000 oder Clearmount f r AEC DAB oder Lichtstark Rot Cat Nr 00 8010 PROBEN PRAPARATION 1 Gewebe in 10 igem neutral gepufferten Formalin fixiert oder anderen Fixiermitteln auf regelm siger Basis benutzen oder gefrorene Gewebesetionen 2 3 4 um Sektionen schneiden und sie auf positiv ges ttigte Objekttr ger legen 3 Uber Nacht bei 37 C trocknen oder f r 2 4 Std bei 58 C VORBEHANDLUNG Hitze Induzierte Epitop Wiedergewinnung HIER wenn erforderlich 1 Objekttr ger deparaffinisieren Gewebesektionen sollten auf Silane Poly L Lysine oder HistoGrip pr pariert werden Cat Nr 00 8050 beschichtete Objekttr ger 2 Objekttr ger mit destilliertem Wasser f r jeweils 2 Min waschen 3 Ein IL Becherglas Pyrex das 500 mL von 0 01 M Citrat Puffer Cat Nr 00 5000 oder EDTA L sung Cat Nr 00 5500 enth lt auf eine heiBe Platte legen Die Pufferl sung erhitzen bis sie kocht Dieser Schritt kann vor der Objekttr ger Deparaffinisierung vo
24. indro Invierta la Jeringa de transferencia Ventana colocando el tap n hacia arriba y el mbolo hacia abajo Luego retire el tap n Haga salir el aire de la Jeringa de transferencia Ventana empujando cuidadosamente el mbolo hacia el tap n Ahora retire el tap n de la punta de la Jeringa de reactivo Ventana Conecte la boquilla de la Jeringa de reactivo Ventana a la boquilla de la Jeringa de transferencia Ventana Dispense anticuerpo de la Jeringa de transferencia Ventana hacia la Jeringa de reactivo Ventana La cantidad m xima que puede contener la Jeringa de reactivo Ventana es 12 5 mL lo cual es suficiente para unas 120 pruebas 9 Coloque la etiqueta Invitrogen adjunta en la Jeringa de reactivo Ventana w cora E El anticuerpo prediluido V Line de Invitrogen est listo para usar con los sistemas de tinci n inmunohistoqu mica Ventana ES NexES y BenchMark Para obtener detalles espec ficos sobre el funcionamiento y la secuencia de procedimientos de los sistemas de tinci n inmunohistoqu mica Ventana ES NexES y BenchMark consulte los manuales del usuario correspondientes y respectivos www invitrogen com Invitrogen Corporation 542 Flynn Rd Camarillo CA 93012 Tel 800 955 6288 E mail techsupport invitrogen com PI 08 4106 Rev 08 08 DCC 08 1089 2007 Invitrogen Corporation All right reserved These products may be covered by one or more Limited Use Label Licenses see th
25. itivo es negativo MARCAS COMERCIALES Cap Plus Clearmount Digest All HistoGrip Histomount y Histostain NBATM PicTureTM y Zymed son marcas comerciales de Zymed Laboratories Inc Ventana EST NexES y BenchMark son marcas comerciales de Ventana Medical Systems Inc 3 x r a P es e e P Los anticuerpos V Line son elaborados exclusivamente por Zymed Laboratories Inc y no implican ni la aprobaci n ni el aval de Ventana Medical ST Systems Inc Las empresas Zymed y Ventana no est n afiliadas asociadas ni relacionadas de manera alguna Para obtener m s informaci n vea la versi n inglesa de la ficha t cnica o vaya a www invitrogen com TIENE ALGUNA PREGUNTA SOBRE ESTE PRODUCTO DIRIJASE A SU DISTRIBUIDOR LOCAL Representante autorizado de IVDD 98 79 EC Invitrogen Ltd Inchinnan Business Park 3 Fountain Drive Paisley PA4 9RF UK www invitrogen com Invitrogen Corporation 542 Flynn Rd Camarillo CA 93012 Tel 800 955 6288 E mail techsupport invitrogen com PI 08 4106 Rev 08 08 DCC 08 1089 2007 Invitrogen Corporation All right reserved These products may be covered by one or more Limited Use Label Licenses see the Invitrogen Catalog or www invitrogen com By use of these products you accept the terms and conditions of all applicable Limited Use Label Licenses e invitrogen CE Souris anti Actin Clone 1 A4 Pour Utilisation de Diagnostiques I
26. n 0 1 sodio azide NaN3 come conservante Immunogeno Peptide sintetico di Actina Isotipo IgG Clone 144 Controllo positivo Leiomiosarcoma Modello di colorazione previsto Citoplasmatica CONSERVAZIONE 2 8 C REQUISITI PER LA COLORAZIONE IMMUNOISTOCHIMICA L Anticorpo diluito V Line di Invitrogen stato sviluppato per l uso con i sistemi di colorazione immunoistochimica Ventana EST NexES e BenchMark insieme ai kit di rilevazione Ventana e gli erogatori di reagente Ventana Invitrogen ha titolato questo anticorpo e lo ha sottoposto ad un controllo della qualit per garantirne la performance coerente ed affidabile La Tabella 1 elenca riassuntivamente i requisiti per l immunocolorazione Tabella 1 Requisiti per Tessuto FFPE Specificazione Recupero di epitopi mediante calore HIER Non Richiesto Digestione enzimatica Non Richiesto Incubazione enzimatica Non Richiesto Incubazione dell anticorpo primario 32 minuti www invitrogen com Invitrogen Corporation 542 Flynn Rd Camarillo CA 93012 Tel 800 955 6288 E mail techsupport invitrogen com PI 08 4106 Rev 08 08 DCC 08 1089 2007 Invitrogen Corporation All right reserved These products may be covered by one or more Limited Use Label Licenses see the Invitrogen Catalog or www invitrogen com By use of these products you accept the terms and conditions of all applicable Limited Use Label Licenses MATERI
27. n Vitro X 08 4106 10 mL Anticorps de pr dilution V Line pr t l emploi Optimis pour les syst mes de coloration immunohistochimique Ventana ES NexES et BenchMark UTILISATION VOULUE Pour Utilisation de Diagnostiques In Vitro L interpr tation doit tre faite par un pathologiste qualifi dans le cadre des ant c dents cliniques du patient et d autres tests de diagnostics REACTIF FOURNI Souris anti Actin est purifi partir de fluide ascite de souris et dilu dans du Phosphate Tamponn Salin PBS pH 7 4 et 1 d albumine de s rum de bovin BSA avec 0 1 azide de sodium NaN comme pr servatif Immunog ne Peptide synth tique actin Isotype IgG Clone 1A4 Contr le Positif L iomyosarcome Mod le de Coloration Attendue Cytoplasmique STOCKAGE 2 8 C MAT RIEL N CESSAIRE POUR LA COLORATION IMMUNOHISTOCHIMIQUE La pr dilution V Line de Invitrogen a t con ue pour tre utilis e avec les syst mes de coloration immunohistochimique Ventana ES NexES et BenchMark en association avec les kits de d tection Ventana et les distributeurs de r actif Ventana Invitrogen a titr et v rifi la qualit de cet anticorps afin de garantir une performance homog ne et fiable Le tableau 1 regroupe le mat riel n cessaire pour l immunocoloration Tableau 1 Mat riel N cessaire pour Tissu FFPE Sp cification Recherche d Epitope de Chaleur par Induction H
28. ng buffer Keep it boiling for 15 minutes 5 After heating remove beaker from the hot plate and allow it to cool down for at least 15 20 minutes at room temperature 6 Rinse slides with PBS Cat No 00 3000 and begin the immunostaining protocol Enzyme Digestion if required 1 See Table 1 for specific enzyme digestion requirement and incubation time LOADING THE USER DEFINED DISPENSER The following procedure is recommended by Invitrogen for filling the Ventana Reagent Dispensers with Invitrogen s V Line Predilute Please refer to the Ventana Operator s Manual for further clarification 1 Remove the plug at the end of the Ventana Transfer Syringe then remove the plunger completely from the barrel and put the plug back on the syringe barrel 2 Appropriately label the Ventana Reagent and Transfer Syringe with the Invitrogen antibody in use 3 Pour the enclosed vial of V Line Predilute 10 mL ready to use antibody into the barrel of the Ventana Transfer Syringe then put the plunger back into the barrel Invert the Ventana Transfer Syringe by positioning the plug upwards and the plunger downward Then remove the plug Displace the air out of the Ventana Transfer Syringe by carefully pushing the plunger towards the plug Now remove the plug from the Ventana Reagent Syringe tip Attach the Ventana Reagent Syringe nozzle to the Ventana Transfer Syringe nozzle Dispense antibody from the Ventana Transfer Syrin
29. ou autres fixatifs de base normale ou des sections gel es de tissu 2 Couper des sections 3 4 um et placer sur des lames charg es positivement 3 S cher toute la nuit 37 C ou pendant 2 4 hrs 58 C PRETRAITEMENT Recherche d Epitope de Chaleur par Induction HIER si n cessaire 1 D paraffiner les lames Les sections de tissu doivent tre mont es sur du silane du poly L Lysine ou du HistoGrip Cat No 00 8050 lames enduites 2 Laver les lames dans 3 bains d eau distill e pendant 2 min chacun 3 Placer un b cher en verre 1L Pyrex contenant 500 mL de 0 01 M tampon de citrate Cat No 00 5000 ou solution d EDTA Cat No 00 5500 sur une plaque chauffante Chauffer la solution de tampon jusqu bullition Cette tape doit tre pr par e ant rieurement au d paraffinage des lames car porter le tampon a bullition peut prendre plusieurs minutes 4 Mettre les lames sur une grille lames et placer les dans le b cher avec le tampon en bullition Laisser bouillir pendant 15 minutes 5 Apr s l chauffement retirer le b cher de la plaque chauffante et laisser le refroidir pour au moins 15 20 minutes temp rature ambiante 6 Rincer les lames dans du PBS Cat No 00 3000 et commencer le protocole d immuno coloration Enzyme de digestion si n cessaire 1 Voir le tableau 1 pour la condition de digestion enzymatique sp cifique et le temps d incubation CHARGEMENT DU DISTRIBUTEUR TEL QUE D
30. r 2 min ciascuno 3 Collocare un beker di vetro Pyrex da IL contenente 500 mL di 0 01 M tampone citrato Cat n 00 5000 o soluzione EDTA Cat No 00 5500 su una piastra scaldante Riscaldare la soluzione tampone sino a quando bolle Questo passaggio pu essere preparato prima della deparaffinizzazione del vetrino poich il tampone pu richiedere diversi minuti per raggiungere il punto di ebollizione 4 Collocare i vetrini in un rack scorrevole e metterli nel beker con tampone bollente Far bollire per 15 minuti 5 Dopo il riscaldamento togliere il beker dalla piastra scaldante e far raffreddare per almeno15 20 minuti a temperatura ambiente 6 Risciacquare i vetrini con PBS Cat n 00 3000 ed iniziare il protocollo di immunocolorazione Digestione Enzimatica se richiesto 1 Per il tempo di incubazione ed i requisiti specifici per la digestione enzimatica consultare la Tabella 1 CARICAMENTO DELL EROGATORE DEFINITO DALL UTENTE Invitrogen raccomanda di osservare la seguente procedura per il riempimento degli erogatori di reagente Ventana con l anticorpo prediluito V Line di Invitrogen Per ulteriori chiarimenti si prega di consultare il Manuale per l utente Ventana 1 Rimuovere il tappo posto sull estremit della siringa di trasferimento Ventana quindi estrarre completamente lo stantuffodal cilindro e rimettere il tappo sul cilindro della siringa 2 Etichettare debitamente la siringa per reagenti e la siringa
31. rbereitet werden da der Puffer mehrere Minuten braucht bevor er kocht 4 Die Objekttr ger in ein Objekttr gergestell setzen und das Becherglas mit dem kochenden Puffer plazieren Es 15 Minuten kochen lassen 5 Nach der Erhitzung das Becherglas von der heissen Platte entfernen und es f r mindestens 15 20 Minuten bei Zimmertemperatur abkiihlen lassen 6 Die Objekttr ger mit PBS Cat Nr 00 3000 sp len und das Immunof rbungs Protokoll beginnen Enzym Verdauung wenn ben tigt 1 Spezifische Enzym Digestionanforderungen und Inkubationszeiten sind Tabelle 1 zu entnehmen F LLEN DES VOM BENUTZER DEFINIERTEN DISPENSERS Invitrogen empfiehlt die folgende Methode zum F llen der Ventana Reagenz Dispenser mit Invitrogen V Line Vorverd nnung Weitere Anweisungen sind dem Ventana Benutzerhandbuch zu entnehmen 1 9 A DA PI 08 4106 Den Verschluss am Ende der Ventana Transferspritze entfernen dann den Kolben ganz aus dem Zylinder herausziehen und den Verschluss wieder auf dem Spritzenzylinder anbringen Die Ventana Reagenz und Transferspritze entsprechend mit dem verwendeten Invitrogen Antik rper kennzeichnen Das beiliegende Fl schchen mit V Line Vorverd nnung 10 mL gebrauchsfertiger Antik rper in den Zylinder der Ventana Transferspritze geben und dann den Kolben wieder in den Zylinder schieben Die Ventana Transferspritze invertieren Dazu den Verschluss nach oben und den Kolben nach unten richten Dann d
32. schen F rbungssystemen Ventana ES NexES und BenchMark in Verbindung mit Ventana Detektionskits und Ventana Reagenz Dispensern entwickelt Invitrogen hat diesen Antik rper titriert und qualit tskontrolliert um dessen konstante und zuverl ssige Leistung zu gew hrleisten In Tabelle 1 sind die Voraussetzungen f r die immunhistochemische F rbung zusammengefasst Tabelle 1 Voraussetzungen fur die FFPE Geweben Angabe Hitze Induzierte Epitop Wiedergewinnung HIER Nicht ben tigt Enzym Verdauung Nicht ben tigt Enzym Inkubation Nicht ben tigt Inkubationsschritte f r den prim ren Antik rper 32 Minuten www invitrogen com Invitrogen Corporation 542 Flynn Rd Camarillo CA 93012 Tel 800 955 6288 E mail techsupport invitrogen com PI 08 4106 Rev 08 08 DCC 08 1089 2007 Invitrogen Corporation All right reserved These products may be covered by one or more Limited Use Label Licenses see the Invitrogen Catalog or www invitrogen com By use of these products you accept the terms and conditions of all applicable Limited Use Label Licenses BEN TIGTE ABER NICHT GELIEFERTE MATERIALIEN Reagens Katalog Nr 1 HistoGrip 00 8050 2 PBS 0 01 M PBS 00 3000 3 Mayer s Haematoxylin 00 8011 4 Zitratpuffer pH 6 0 falls f r die durch W rmeeinwirkung induzierte Wiedergewinnung der Epitope erforderlich 00 5000 5 EDTA L sung falls f r die durch W rmeeinwirkung induzierte W
33. t 100 Ensuite dans du xyl ne et xyl ne Ne pas d shydrater lorsque l AEC est utilis 2 minutes chacun Appliquer le couvre lames avec une solution de fixation Histomount utiliser la solution de fixation GVA lorsque l AEC est utilis Laisser s cher avant de visualiser 10 minutes DEPANNAGE Probl me Solution L immunocoloration du t moin Examiner d autres t moins positifs pr par s au cours de la m me analyse afin de d terminer si elle positif est faible est due l anticorps primaire ou un des r actifs communs secondaires V rifier que la lame comprend la bonne tiquette de code barres de Ventana Si elle est bien Le t moin positif est n gatif tiquet e v rifier d autres t moins positifs pr par s au cours de la m me analyse afin de d terminer si c est d l anticorps primaire ou un des r actifs communs secondaires MARQUE DE FABRIQUE Cap Plus Clearmount Digest All HistoGrip Histomount et Histostain NBATM PicTure et Zymed sont des marques des Zymed Laboratories Inc Ventana ES NexES et BenchMark sont des marques d pos es de Ventana Medical Systems Inc Les anticorps V Line sont d velopp s exclusivement par Zymed Laboratories Inc et n insinuent pas une approbation ou un soutien de la part de Ventana Medical Systems Inc Zymed et Ventana ne sont en aucune fa on affili es ou associ es
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