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A. Avogadro. Indiana University Southeast.

This has proven highly successful for the demon- read by light microscopic observation 2.5 mg cialis overnight delivery erectile dysfunction caused by high cholesterol. Tissue sections preserved in Immunodiagnosis involves the use of antibody assays cheap cialis 20 mg without a prescription erectile dysfunction under 35, paraffn are frst treated with xylene purchase cialis 2.5 mg visa erectile dysfunction blood pressure, and after deparaffni- immunocytochemistry, the identifcation of lymphocyte zation they are exposed to a hydrogen peroxide solution that markers, and other techniques to diagnose infectious dis- destroys the endogenous peroxidase activity in tissue. This antigen is expressed by 84% of invasive ductal breast carci- so-called linking antibody will combine with any primary noma and 85 to 90% of colon, pancreatic, gastric, esophageal, rabbit antibody in the tissue. It is added in excess, which lung (non-small cell), ovarian, and endometrial adenocarci- will result in one of its antigen-binding sites remaining free. Lung adeno- formalin resistant, permitting the detection of ovarian cancer carcinoma rarely stains with this antibody. Normal apocrine sweat nal antibody is specifc for a regulatory protein present in glands, eccrine glands (variable), minor salivary glands, smooth muscle and selected other tissues. It interacts with bronchial glands, metaplastic breast epithelium, benign sweat actin, myosin, tropomyosin and calmodulin. It participates in smooth muscle contrac- with greater reliability than nonspecifc stains (e. It may also be positive in malignant myothepi- Antibroad-spectrum cytokeratin is a mouse monoclonal thelioma, pleomorphic adenoma of the salivary glands, and antibody that may be used to identify cells of normal and angiomatoid malignant fbrous histiocytoma. The pankeratin cocktail recognizes was later shown to be present in certain other conditions as most of the acidic and all of the basic cytokeratins, making it well. This antibody binds specifcally to antigens located cervical, bladder, medullarythyroid, and prostatic carcinoma. The antibody is used to qualitatively stain cytok- noma, lymphoproliferative disease, and smokers. In anaplastic tumors, the per- to the level immediately following surgery, this may signify centage of tumor cells showing cytokeratin reactivity may be metastases. Unexpected antigen expression or loss of peptide chain with one variable region at the amino terminus expression may occur, especially in neoplasms. It is detected with a mouse of any staining or its absence must be complemented by mor- monoclonal antibody directed against a complex glycoprotein phological studies and evaluation of proper controls. This reagent may be used to aid in the identifcation of cells Anti-high molecular weight human cytokeratin antibod- of epithelial lineage. The antibody is intended for qualitative ies are mouse monoclonal antibodies that identify keratins of staining in sections of formalin-fxed, paraffn-embedded tis- approximately 66 kDa and 57 kDa in extracts of the stratum sue. The antibody labels squamous, ductal, and other in the plasma membrane and cytoplasmic regions of normal complex epithelia. Unexpected antigen expression or loss of neoplasms and variably with those derived from simple epithe- expression may occur, especially in neoplasms. Consistently positive are squamous cell carcinomas and stromal elements surround heavily stained tissue and/or cells ductal carcinomas, most notably those of the breast, pancreas, which show immunoreactivity. Clinical interpretation of any bile duct, and salivary gland; transitional cell carcinomas of staining or its absence must be complemented by morphologi- the bladder and nasopharynx; and thymomas and epithelioid cal studies and evaluation of proper controls. Antibodies to this molecule melanomas, neural tumors, and neuroendocrine tumors are confrm its presence and reveal the morphological appearance unreactive. Normal tissue stains with this antibody in a fashion consistent with the sites of mesenchymal elements and Cytokeratin (34betaE12), mouse: Anticytokeratin (34beta epithelial basal laminae. This antibody recognizes cytokeratins 1,5,10, and 14 neoplasma, hemangiopericytoma, angiosarcoma, and epithe- that are found in complex epithelia. Cytokeratin 34betaE12 liod hemangioendothelioma can be revealed by this antibody shows no reactivity with hepatocytes, pancreatic acinar Diagnostic Immunohistochemistry 871 cells, proximal renal tubes, or endometrial glands; there has from other sites were not positive using the antibody to been no reactivity with cells derived from simple epithe- cytokeratin 20, e. Mesenchymal tumors, lymphomas, melanomas, neural and endometrium, and nonmucinous tumors of the ovary. There was a lack of positivity in ful in distinguishing prostatic adenocarcinoma from hyper- small-cell lung carcinomas and in intestinal and pancreatic plasia of the prostate. Anti-low molecular weight cytokeratin is a mouse mono- Antihuman cytokeratin 7 antibody (Figure 29. It may be used to aid in the identifcation of cells tin intermediate flament protein identifed as cytokeratin 7, of epithelial lineage. The antibodies are intended for qualitative a basic cytokeratin found in most glandular epithelia and staining in sections of formalin-fxed paraffn-embedded tis- in transitional epithelia. Antikeratin primary antibody specifcally binds to antigens number of epithelial cell types including many ductal and located in the cytoplasmic regions of normal epithelial cells. In general, the antibody does not react Unexpected antigen expression or loss of expression may occur, with stratifed squamous epithelia but is reactive with tran- especially in neoplasms. The antibody reacts rounding heavily stained tissue and or cells will show apparent with many benign and malignant epithelial lesions. The clinical interpretation of any staining, or 7 is expressed in specifc subtypes of adenocarcinomas from its absence, must be complemented by morphological studies ovary, breast, and lung, whereas carcinomas from the gastro- and evaluation of proper controls. Transitional cell carcinomas express keratin 7, whereas prostate cancer is generally nega- Antihuman cytokeratin-20 monoclonal antibody (Fig- tive. In cytological specimens, the foveolar epithelium, a number of endocrine cells of the upper antibody permits ovarian carcinoma to be distinguished from portions of the pyloric glands, as well as with the urothe- colon carcinoma. The antibody has been tested on a series of carcinomas including primary and Cytokeratin 7 (K72), mouse: Anticytokeratin 7 (K72) mouse metastatic lesions. There is a marked difference in expression monoclonal antibody reacts with proteins that are found in of cytokeratin 20 among various carcinoma types. Neoplasia most ductal, glandular, and transitional epithelium of the expressing cytokeratin 20 are derived from normal epithe- urinary tract and bile duct epithelial cells. Colorectal carcinomas con- distinguishes between lung and breast epithelium that stain sistently express cytokeratin 20, whereas adenocarcinomas positive, and colon and prostate epithelial cells that are nega- of the stomach express cytokeratin 20 to a lesser degree. This antibody also reacts with many benign and malig- Adenocarcinomas of the gall bladder and bile ducts, ductal nant epithelial lesions, e. Transitional cell carcinomas are positive tumors, and transitional-cell carcinomas have been found to and prostate cancer is negative. It stains positively in ity among the 19 catalogued human epidermal keratins and glandular epithelium as well as adenocarcinomas of the lung produces positive staining in virtually all epithelia. Nineteen different marker that identifes, by immunoperoxidase staining, most molecular forms of cytokeratin have been identifed in both epithelial cells and tumors derived from them, such as breast normal and malignant epithelial cell lines. Thus, it must be used in conjunction with other markers tant markers for classifying carcinomas (tumors of epithelial in tumor identifcation and/or classifcation. The identifcation of cytokeratin has mouse monoclonal antibody directed against a mucin epitope gained increasing importance in immunopathology. Epithelial membrane monoclonal antibody detects E-Cadherin, an adhesion protein antigen is widely distributed in epithelial tissues and tumors arising from them. Normal glandular epithelium and tis- sue from nonneoplastic diseases stain in lumen membranes and cytoplasm. Malignant neoplasms of glandular epithe- lium frequently show a change in pattern with the appear- ance of adjacent cell membrane staining. Estrogen/progesterone receptor protein: Monoclonal antibodies against estrogen receptor protein and against progesterone receptor protein permit identifcation of tumor cells by their preferential immunoperoxidase staining for these markers, whereas stromal cells remain unstained. This method is claimed by some to be superior to cytosol assays in evaluating the clinical response to hormones. Both mouse and rabbit monoclonal antibodies have been used for immunohistochemical staining and anti-Her2/ Neu (c-erb-B-2) preparations.

The tendon of the flexor carpi ulnaris tendon will be evident closest to the little finger cheap cialis 10 mg online erectile dysfunction ed treatment. A high-frequency linear ultrasound transducer is placed in a transverse position over the tendon and an ultrasound survey scan is taken (Fig order cialis toronto erectile dysfunction treatment in lucknow. The tendon of the flexor carpi ulnaris tendon should appear on the ulnar side of the ulnar nerve and the ulnar artery buy cialis 20 mg without prescription impotence back pain. The ulnar nerve appears as a bundle of hyperechoic nerve fibers surrounded by a slightly more hyperechoic neural sheath lying beneath the flexor retinaculum with the ulnar artery just radial to it (Fig. The median nerve can be distinguished from the flexor tendons by simply having the patient flex and extend their fingers and observing the movement for the tendons. The flexor tendons will also exhibit the property of anisotropy with the tipping of the ultrasound transducer back and forth over the tendons. The flexor carpi ulnaris will be the most ulnar and superficial of the superficial flexor tendons and can be seen to lie just above the dome-shaped pisiform (Fig. There may be significant effusion surrounding the tendon which will appear on transverse ultrasound imaging as a hypoechoic ring around the tendon. If there is a question as to whether the tendon is the flexor carpi ulnaris tendon, the ultrasound transducer can be turned to the longitudinal plane and the tendon can be followed distally to its insertion on the pisiform (Fig. Color Doppler may identify hyperemia of the musculotendinous unit and may also be useful in helping identify the ulnar artery if the anatomy is not clear (Fig. After the musculotendinous unit is identified as it passes under the flexor retinaculum, the tendon is evaluated for tendinopathy, tendinitis, tear, extrinsic compression, and rupture (Fig. A: Identification of the flexor carpi ulnaris tendon is facilitated by having the patient forcibly flex his or her wrist. B: Proper ultrasound transducer placement for ultrasound-guided injection for flexor carpi ulnaris tendinitis. Transverse ultrasound view of the flexor carpi ulnaris tendon at the wrist and its relationship to the ulnar nerve and artery. Longitudinal ultrasound view of the flexor carpi ulnaris tendon demontraing its insertion on the trapezium. Color Doppler can be useful in helping identify the ulnar artery which lies just radial to the ulnar nerve and the flexor carpi ulnaris tendon. A: Clinical appearance of severed extensor carpi ulnaris tendon following a cutting injury. An erythematous and tender mass located 2 cm distal to the scar that was left after the repair of a laceration following a cutting injury. Neglected ruptured flexor carpi ulnaris tendon mimics a soft tissue tumor in the wrist. Physical examination combined with judicious use of ultrasound, magnetic resonance imaging, and radiography will help delineate the cause of ulnar-sided wrist pain. The function of the abductor pollicis longus and extensor pollicis brevis muscles is radial abduction of the thumb. The radial artery and the superficial branch of the radial nerve are in proximity to the site for injection to treat de Quervain tenosynovitis and these structures may be traumatized if the needle is placed too medially (Fig. De Quervain tenosynovitis, which is also known as mommy’s thumb or wrist, is caused by inflammation of the tendons and tendon sheaths of the abductor pollicis longus and extensor pollicis brevis muscles. The relationship between the radial styloid, the tendons, and tendon sheaths of the abductor pollicis longus and extensor pollicis brevis muscles, and the radial artery and superficial branch of the radial nerve. This painful condition is named for Swiss surgeon Fritz de Quervain who first described this constellation of symptoms and their cause in 1895. The result of repetitive high-torque twisting motions of the wrist and occasionally as a result of direct trauma to the tendons of the abductor pollicis longus and extensor pollicis brevis at the level of the radial styloid process, de Quervain tenosynovitis can cause significant pain and functional disability if not promptly treated. On rare occasions, de Quervain tenosynovitis can develop without antecedent trauma, especially in the parturient and this setting is often referred to as mommy’s thumb or wrist. The symptoms of de Quervain tenosynovitis are the result of inflammation and edema of the tendons and tendon sheath of the abductor pollicis longus and extensor pollicis brevis muscles at the level of the radial styloid process (Fig. If untreated, a 495 thickening of the tendons and tendon sheath may occur, resulting in a constrictive tenosynovitis. In some patients, a triggering phenomenon of the thumb may occur as a result of the thickened tendon locking or catching in the constricted tendon sheath. Arthritis and gout of the first metacarpal joint also may coexist with and exacerbate the pain and disability of de Quervain tenosynovitis. Transverse ultrasound image of the first dorsal compartment tendons (abductor pollicis longus and extensor pollicis brevis) showing tenosynovitis. Activities associated with the development of de Quervain tenosynovitis include repetitive hand shaking, scooping ice cream, or using a screw driver. The pain of de Quervain tenosynovitis is sharp and constant and is exacerbated by any activities requiring active pinching of the thumb or ulnar deviation of the wrist. The pain is localized to the area over the radial styloid process and is associated with increasing functional disability if the inflammatory process remains untreated. On physical examination, there is tenderness and swelling over the tendons and tendon sheaths along the distal radius, with point tenderness over the radial styloid. A creaking tendon sign may be noted with flexion and extension of the thumb and triggering of the thumb may occur. Patients with de Quervain tenosynovitis demonstrate a positive Finkelstein test (Fig. The Finkelstein test is performed by stabilizing the patient’s forearm, having the patient fully flex his or her thumb into the palm, and then actively forcing the wrist toward the ulna. Patients suffering from de Quervain tenosynovitis will exhibit a positive Finkelstein test. Plain radiographs of the wrist are indicated in all patients suspected of suffering from de Quervain tenosynovitis to rule out occult bony pathology and to identify calcific tendinitis. Based on the patient’s clinical presentation, additional testing may be indicated, including complete blood cell count, uric acid, sedimentation rate, and antinuclear antibody testing. Magnetic resonance imaging and ultrasound imaging of the wrist is indicated to assess the status of the abductor pollicis longus and extensor pollicis brevis tendons and tendon sheath as well as to identify other occult pathology including arthritis and gout involving the first metacarpal joint (Fig. Longitudinal ultrasound image of De Quervain tenosynovitis in a volleyball player shows thickening of the extensor carpi radialis. With the patient in the above position, the radial styloid process and the abductor pollicis longus and extensor pollicis brevis tendons at that level are identified by palpation. Identification of the tendons is facilitated by having the patient radially deviate the wrist against the examiner’s resistance (Fig. At the level of the radial styloid a high-frequency linear ultrasound transducer is placed in a transverse position over the abductor pollicis longus and extensor pollicis brevis tendons and an ultrasound survey scan is taken (Figs. Color Doppler may aid in identification of the radial artery and help separate it with the superficial radial nerve which lies just radial to the radial artery (Fig. The tendons will appear as the hyperechoic “hole” in the hypoechoic tendon sheath. However, in a small number of patients, the tendon sheath will appear to travel through separate subcompartments divided by a subcompartmental septum (Fig.

A similar procedure is used in detect- ing antibodies against intercellular substance antigens in the the indirect fuorescence antibody technique is a method serum of patients with pemphigus vulgaris buy cheap cialis on-line impotence grounds for divorce in tn. The indirect test has a greater sensitivity than the direct fuorescence antibody method (Figure 28 buy cialis paypal men's health erectile dysfunction causes. It is often employs antibodies buy cialis 20 mg with visa impotence high blood pressure, either polyclonal or monoclonal, labeled referred to as the sandwich or double-layer method. When T or B lymphocytes containing Quin-2 are activated, their fuorescence intensity rises, implying an elevation of free Ca++ in the cytosol. Tissue Ferritin is an iron-containing protein that is electron dense and serves as a source of stored iron until it is needed for the synthesis of hemoglobulin. Ferritin’s electron-dense quality makes it useful to label antibodies or antigens to be identifed or localized in electron microscopic preparations. Antibodies may be labeled with ferritin by use The binding of ferritin particles to the surface of the erythrocyte of a cross-linking reagent such as toluene-2,4-diisocyanate. Immunohistochemistry is a method to detect antigens in tissues that employs an enzyme-linked antibody specifc for Immunoelectron microscopy refers, traditionally, to the antigen. The enzyme degrades a colorless substrate to a col- use of antibodies labeled with ferritin to study the ultrastruc- ored insoluble substance that precipitates where the antibody ture of subcellular organelles and, more recently, the use of and, therefore, the antigen are located. Identifcation of the immunogold labeling and related procedures for the identif- site of the colored precipitate and the antigen in the tissue cation and localization of antigens by electron microscopy. Diagnostic pathology services routinely offer approximately 100 anti- Theimmunoferritin method (Figure 28. Immunoglobulin may be conjugated with ferritin, an electron-dense marker, without altering its immunological reactivity. These ferritin- labeled antibodies localize molecules of antigen in the sub- cellular areas. Electron-dense ferritin permits visualization of antibody binding to homologous antigen in cells and tissues by electron microscopy. In addition to ferritin, horseradish- peroxidase-labeled antibodies may also be adapted for use in immunoelectron microscopy. Sections are incubated with primary antibody and followed by treatment with colloidal gold-labeled anti-IgG antibody. Electron-dense particles are localized at sites of antigen– antibody interactions. The immunoperoxidase tech- than that of most other methods when examined in a bright nique permits the demonstration of antigens in various types feld or in conjunction with polarized light. This method has certain advan- successfully stains tissue sections from paraffn wax, resin, tages that include (1) the use of conventional light micro- or cryostat preparations. It is also effective for cell suspen- scopy, (2) the stained preparations may be kept permanently, sions or smears, cytospin preparations, cell cultures, or tissue (3) the method may be adapted for use with electron micro- sections. Both 1- and 5-nm gold conjugates are used for light scopy of tissues, and (4) counterstains may be employed. The 1-nm particles are advantageous in studies disadvantages include the following: (1) the demonstration of cell penetration. In immunogold silver staining, primary of relatively minute positively staining areas is limited by antibody is incubated with tissues or cells to localize anti- the light microscope’s resolution, (2) endogenous peroxidase gens that are identifed with gold-labeled secondary antibod- may not have been completely eliminated from the tissue ies and silver enhanced. Because avidin has Rabbit + such an extraordinarily high affnity for biotin, the binding of primary avidin to biotin is essentially irreversible. Antigen retrieval is a novel method for the rescue of anti- + gens from formalin-fxed paraffn-embedded tissue. It con- sists of heating sections in a microwave oven or in a pressure cooker in the presence of an antigen retrieval solution. This technique increases staining intensity and reduces background staining of many important markers in formalin-fxed tissue. Its use helps overcome false-negative staining of overfxed tissue, expands the range of antibod- ies useful for routinely processed tissue, and increases the usefulness of archival materials for retrospective studies. In addition to microwave heating, the pH of the antigen retrieval solution is an important cofactor for some antigens. These include a citrate-based neutral pH solution, Tris-based high pH solution, and a glycine-based low pH solution. This has proven sites for biotin) highly successful for the demonstration of antigens in par- Biotin affn-embedded tissues as an aid in surgical pathologic diag- Peroxidase nosis. Tissue sections preserved in paraffn are frst treated with xylene, and after deparaffnization they are exposed to Development in chromogenic hydrogen donor and hydrogen a hydrogen peroxide solution which destroys the endogenous peroxide. The sections are next incubated brown granular deposit depending upon the chromogenic hydrogen donor used. Thereafter, the primary rabbit antibody against the Immunological Methods and Molecular Techniques 839 antigen to be identifed is reacted with the tissue section. Primary antibody that is unbound is removed by rinsing the sections which are then covered with swine antibody against rabbit immunoglobulin. This so-called linking antibody will combine with any primary rabbit antibody in the tissue. It is added in excess, which will result in one of its antigen- binding sites remaining free. The tis- sue sections can then be counterstained with hematoxylin or other suitable dye, covered with mounting medium and cover slips, and read by conventional light microscopy. For human primary antibody, an additional step must link the human anti- body into the rabbit sandwich technique. Paraffn-embedded tissue sections are frst treated with xylene, and after deparaf- fnization, they are exposed to hydrogen peroxide to destroy the endogenous peroxidase. Sections are next incubated with normal sheep serum to suppress nonspecifc binding of immu- noglobulin to tissue collagen. Primary rabbit antibody against the antigen to be identifed is combined with the tissue section. Unbound primary antibody is removed by rinsing the sec- tions which are then covered with sheep antibody against rab- bit immunoglobulin. This linking antibody will combine with any primary rabbit antibody in the tissue. It is added in excess, which results in one of its antigen-binding sites remaining free. Biotin–avidin system: Avidin is an egg white-derived glycoprotein with an extraordinarily high affnity (affnity constant > 1015M–1) for biotin. Streptavidin is similar in prop- erties to avidin but has a lower affnity for biotin. Many biotin molecules can be coupled to a protein, enabling the biotiny- lated protein to bind more than one molecule of avidin. If biotinylation is performed under gentle conditions, the bio- logical activity of the protein can be preserved. This system has proven particularly useful in the brown granular deposit depending upon the chromogenic hydrogen donor used. Pancreatic islet cell hormones: Immunoperoxidase stain- area of reddish-brown pigment.