protocols-10
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钟意阿满
怎么正确的使用SQLNET.ALLOWED
根据自己的实验目的和实验对象而异.提供一个protocol,可参考:。
CD79a (B-cell) Immunohistochemical Staining Protocol。
Description: Anti-CD79a is to identify B-cell linage (but not in myeloid or T cell lines) in formalin-fixed, paraffin embedded normal and neoplastic tissues using immunohistochemical (IHC) test methods. The CD79á polypeptide appears early in B-cell maturation at the pre-B-cell stage, and persists until the plasma cell stage, where it is found as an intracellular component. Anti-CD79á can be used for the immunohistochemical identification of cells of the B-cell lineage.。
Fixation: Formalin-fixed, paraffin embedded sections and acetone-fixed cryostat sections and fixed cell smears.。
Positive Controls: Tonsil, thymus, spleen.。
Solutions and Reagents:。
Primary Antibody:。
Mouse Anti-Human CD79a, B-Cell (Clone JCB117) (Dakocytomation, Cat# M7050). Optimal dilution 1:100. Species Reactivity: Human (refer to antibody datasheet for more information).。
Secondary Antibody:。
Horse Anti-Mouse IgG (H+L), biotinylated (Vector Laboratories, Cat# BA-2000). Optimal dilution 1:500.。
Detection Reagent:。
HRP-Streptavidin (Vector Laboratories, Cat# SA-5004). Optimal dilution 1:500。
Procedure:
1. Sections to distilled water (refer to general IHC protocols as needed).。
2. Epitope Retrieval: Use Tris-EDTA Buffer Epitope Retrieval Method. Briefly, pre-heat steamer or water bath with staining dish containing Tris-EDTA Buffer (pH 9.0) until temperature reaches 95-100 °C. Immerse slides in the staining dish and place the lid loosely on the staining dish. Incubate for 20 minutes and turn off the steamer or water bath. Place the staining dish at room temperature and allow the slides to cool for 20 minutes.。
3. Rinse sections in 2 changes of washing buffer, 2 minutes each.。
4. Serum Blocking: incubate sections with normal horse serum blocking solution for 30 minutes to block non-specific binding of immunoglobulin.。
5. Primary Antibody: incubate sections with Mouse Anti-Human CD79a, B-Cell (Clone JCB117) (Dakocytomation, Cat# M7050) diluted 1:100 in primary antibody dilution buffer for 1 hour at room temperature.。
6. Rinse in washing buffer for 2x2min.。
7. Peroxidase Blocking: incubate sections in peroxidase blocking solution for 10 minutes to block endogenous peroxidase activity.。
8. Rinse in washing buffer for 3x2 min.。
9. Secondary Antibody: incubate sections with biotinylated Horse Anti-Mouse IgG diluted in secondary antibody dilution buffer for 30 minutes at room temperature.。
10. Rinse in washing buffer for 3x2min.。
11. Detection: incubate sections with HRP-Streptavidin diluted in HRP-streptavidin dilution buffer for 30 minutes at room temperature.。
12. Rinse in washing buffer for 3x2min.。
13. Chromogen/Substrate: incubate sections in DAB peroxidase substrate solution for 5-10 minutes.。
14. Rinse in distilled water briefly.。
15. Counterstain with Gill's hematoxylin solution or Mayer's hematoxylin solution if desired.。
16. Rinse in running tap water for 5 minutes.。
17. Dehydrate through 95% ethanol for 2 minutes, 100% ethanol for 2x3min.。
18. Clear in xylene for 2x3min.。
19. Coverslip with permanent mounting medium.。
Results:
Staining pattern: Cytoplasmic (Search Images)。
Notes:
1. Avidin/Biotin Blocking may be needed to block endogenous biotin activity for certain tissues such as kidney, liver, prostate, colon and gut, which may contain endogenous biotin.。
2. For frozen sections, snap frozen fresh tissues in isopentane pre-cooled in liquid nitrogen, embedded in OCT compound in cryomolds. Cut 4-8 um cryostat sections and mount on superfrost plus slides. Store slides at - 80°C until needed. Before staining, air dry slides at room temperature for 30 minutes and fix in ice-cold acetone for 5 minutes. Air dry for another 30 minutes. Then start from step 3 for routine immunostaining.。
回答采摘自文献，仅供参考哈
抱起亚轩找小葵
正确理解sqlnet.ora参数SQLNET.ALLOWED_LOGON_VERSION是怎么工作的。
解决方案：
关于sqlnet.ora参数SQLNET.ALLOWED_LOGON_VERSION以及该参数怎么影响oracle client连接oracle database，存在通常上的误解。
该参数实际上指定了一个client被允许使用的AUTHENTICATION PROTOCOL (for example SHA-1)，而不是该client的实际版本（version）
举例：
SQLNET.ALLOWED_LOGON_VERSION=10。
即使该参数值暗示oracle database version 10,但是内部检查实际上是针对authentication protocol 'SHA-1'.。
Even though the parameter value implies Oracle version 10 the internal check is really against the authentication protocol 'SHA-1'.。
在更早的Oracle version中，这是一对一的关系（one-on-one relation），比如：一个特定的Oracle client版本使用特定的authentication protocol。
从Oracle 10g开始，不再是一对一的关系：Oracle 10g和11g 都使用SHA-1 protocol。 Oracle 12g(译者注：原文就是12g)将会使用SHA-2 protocol。
这意味着：通过使用该参数，没有方法防止10g的client连接 并仅仅允许11g的client连接，因为10g和11g使用相同的SHA-1 protocol。这也意味着这两个客户版本是一样安全的---通过单独尝试防止10g client的连接，不会在安全上获益。
该参数在文档上是含混不清的(is unclear),so Doc bug 11845659 - sqlnet.allowed_logon_version needs clearer documentation has been created to fix the description of this parameter.。
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