Species identification and antifungal susceptibility profile of candida isolates from ICU patients in Sohag University hospital, Upper Egypt

Background: Candida species are among the most common fungal pathogens in ICU patients. Candida albicans was the predominant species, but a shift toward non-albicans Candida species has been recently observed

Objectives: To detect the prevalence of different Candida species and determine their antifungal susceptibility profile in ICU patients using phenotypic methods, the Vitek 2 system compared with CHROMagar Candida agar and a genotypic method; PCR-RFLP

Methodology: Various clinical samples were collected from 248 ICU patients in Sohag University Hospital from the period between September 2014 and May 2015. Samples were cultured on CHROMagar Candida agar. Results were compared with those of Vitek 2 system and confirmed by PCR- RFLP method and antifungal susceptibility profiles were analyzed by disc diffusion and Vitek 2 antifungal susceptibility tests

Results: The study revealed an overall isolation rate of Candida species among ICU patients was 29% by PCR-RFLP. Candida albicans was the most frequent species isolated [40.3%]. Non- albicans Candida species including Candida tropicalis [22.2%], Candida glabrata [18%], Candida krusei [12.5%], C. parapsilosis [4.2%], C. dubliniensis [1.4%] and Candida guilliermondii [1.4%] were also isolated. The sensitivity of vitek 2 with regard to correct identification of Candida species was 96%; the specificity was 100%, also CHROMagar Candida agar enable the correct identification with sensitivity 89%, specificity 100%. Vitek 2 antifungal susceptibility tests results were found to be an accurate method as it was compared with the disc diffusion method for fluconazole, voriconazole and amphotracin B

Conclusion: CHROMagar Candida agar supported by Vitek 2 system is a valuable method for identification of common Candida species, these methods are easy to interpret and give rapid results in comparison with the expensive PCR-RFLP method. Although amphotericin B and fluconazole are widely used in clinical practice, there was no evidence of enhanced resistance. Moreover, voriconazole could be used in treatment of fluconazole-resistant Candida species

Evaluación de la actividad antifúngica de extractos liquénicos e identificación de sus metabolitos / Evaluation of the fungicide activity from lichens extracts and identification their metabolites

Los líquenes son los hongos que establecen una relación simbiótica con un alga o cianobacteria. En esta simbiosis se producen por parte del hongo, una serie de metabolitos secundarios conocidos como sustancias liquénicas; las cuales presentan una marcada actividad antibiótica. En Cuba no se tienen antecedentes sobre estudios de metabolitos liquénicos por lo que se propone; evaluar el efecto fungicida de extractos liquénicos producidos por especies cubanas así como identificar sus metabolitos. Se emplearon líquenes de diferentes zonas del país (Parmotrema dilatatum, P. tinctorum, P. praesorediosum P. cristiferum, Ramalina americana, Cladonia ceratophylla y Cladonia portentosa spp. pacífica), a los cuales se les extrajo con acetona, las sustancias liquénicas almacenadas en el talo. Los extractos fueron probados contra los hongos fitopatógenos Rhizoctonia solani y Phythophtora nicotianae; por el método de envenenamiento del medio de cultivo agar papa dextrosa a concentraciones de: 0,01 por ciento; 0,03 por ciento y 0,07 por ciento. Se utilizó un control negativo de dimetilsufóxido al 0,07 por ciento y se determinaron los porcentajes de inhibición, cuyos resultados fueron analizados estadísticamente. Los metabolitos secundarios presentes en los extractos se identificaron por cromatografía de capa fina (TLC). Exceptuando el extracto liquénico de P. cristiferum, todos los demás mostraron más de un 50 por ciento de inhibición del crecimiento de ambos hongos a la concentración de 0,07 por ciento, mientras que a las restantes concentraciones los valores fueron variados con diferencias significativas con respecto al control. Se lograron identificar tres metabolitos liquénicos: metil 2‘-O- metilmicrofilinato, 4-O-Demetilmicrofilinico y el ácido ramaniloico.

Lichens are fungi that establish a symbiotic relationship with an alga or cyanobacterium. This symbiosis produced by the fungus, a series of secondary metabolites known as lichen substances; which show a strong antibiotic activity. In Cuba there is no background on the studies above lichen metabolites so it is proposed; evaluate the fungicidal activity of lichen extracts produced by Cuban species and to identify metabolites. Lichens from different areas of the country (Parmotrema dilatatum, P. tinctorum, P.praesorediosum, P. cristiferum, Ramalina americana, Cladonia ceratophylla and Cladonia portentosa spp. pacífica), to which it extracted with acetone, the lichen substances stored in tallus. The extracts were tested against the fungal pathogens Rhizoctonia solani and Phythophtora nicotianae; poisoning by the method of culture medium potato dextrose agar at concentrations of 0.01 percent; 0.03 percent and 0.07 percent. A negative control to 0.07 percent dimethylsulfoxide was used and the percentage of inhibition, the results were analyzed statistically determined. Secondary metabolites present in the extracts were identified by thin layer chromatography (TLC). Except P. cristiferum lichen extract, all others showed more than 50 percent growth inhibition of both fungi at concentration of 0.07 percent, while the remaining concentrations were varied values with significant differences from the control. Was made to identify three lichen metabolites metilmicrofilinato 2‘ methyl-O, 4-ODemetilmicrofilinico and ramaniloico acid.

Study on chemical constituents from branches and leaves of Polyalthia nemoralis / 中国中药杂志

<p><b>OBJECTIVE</b>To investigate the chemical constituents of the branches and leaves of Polyalthia nemoralis.</p><p><b>METHOD</b>The compounds were isolated and purified by silica gel, macroporous adsorption resin and Sephadex LH-20 column chromatographic methods. Their chemical structures were elucidated on the basis of physicochemical properties and spectral data.</p><p><b>RESULT</b>Fourteen compounds were isolated and identified as syringic acid (1), 3-methoxy-4-hydroxycinnamic acid (2), vanillic acid (3), 4-hydroxybenzoic acid (4), mauritianin (5), (+)-xylopinidine (6), (+)-oblongine(7), (+)-tembetarine (8), eythritol (9), D-mannitol (10), ethyl-beta-D-glucopyranoside (11), (+)-magnoflorine (12), stepharanine (13), (2S, 4R)-4-hydroxy-2-piperidine-carboxylic acid (14), respectively.</p><p><b>CONCLUSION</b>All the compounds were isolated from the genus Polyalthia for the first time; compounds 6 and 13 showed inhibitation activities against multi tumor cell lines.</p>

Chemical constituents of Ammopiptanthus mongolicus / 中国中药杂志

<p><b>OBJECTIVE</b>To study the chemical constituents of aerial parts of Ammopiptanthus mongolicus.</p><p><b>METHOD</b>Isolation and purification were carried out on silica gel, Sephadex LH-20 and HPLC column chromatography. The structures of the compounds were identified by physico-chemical properties and spectral analysis.</p><p><b>RESULT</b>Nine compounds were isolated and identified as (+)-maackiain (1), brevifolin (2), 7-hydroxy-4'-methoxy isoflavanone (3), daidzein 4',7-diglucoside (4), genistein 4', 7-di-O-beta-D-glucoside (5), isolupalbigenin (6), ononin (7), beta-sitosterol (8), beta-daucosterol (9).</p><p><b>CONCLUSION</b>Compounds 2, 4 - 6 were obtained from the genus Ammopiptanthus for the first time.</p>

Isolation and characterization of a serine proteinase with thrombin-like activity from the venom of the snake Bothrops asper

A serine proteinase with thrombin-like activity was isolated from the venom of the Central American pit viper Bothrops asper. Isolation was performed by a combination of affinity chromatography on aminobenzamidine-Sepharose and ion-exchange chromatography on DEAE-Sepharose. The enzyme accounts for approximately 0.13 percent of the venom dry weight and has a molecular mass of 32 kDa as determined by SDS-PAGE, and of 27 kDa as determined by MALDI-TOF mass spectrometry. Its partial amino acid sequence shows high identity with snake venom serine proteinases and a complete identity with a cDNA clone previously sequenced from this species. The N-terminal sequence of the enzyme is VIGGDECNINEHRSLVVLFXSSGFL CAGTLVQDEWVLTAANCDSKNFQ. The enzyme induces clotting of plasma (minimum coagulant dose = 4.1 µg) and fibrinogen (minimum coagulant dose = 4.2 µg) in vitro, and promotes defibrin(ogen)ation in vivo (minimum defibrin(ogen)ating dose = 1.0 µg). In addition, when injected intravenously in mice at doses of 5 and 10 µg, it induces a series of behavioral changes, i.e., loss of the righting reflex, opisthotonus, and intermittent rotations over the long axis of the body, which closely resemble the `gyroxin-like' effect induced by other thrombin-like enzymes from snake venoms.

Purification of elastase-like chymotrypsin from cardamom shoot and Capsule borer [corrected]

An elastase-like chymotrypsin was purified by aprotinin-agarose affinity chromatography from the midgut extract of cardamom shoot and capsule borer, Conogethes punctiferalis. The purified enzyme had a Vmax of 687.6 +/- 22.1 nmole pNA released/min/mg protein, Km of 0.168 +/- 0.012 mM with SAAPLpNA as substrate and gave a single band on SDS-PAGE with a molecular mass of 72.1 kDa. Casein zymogram revealed one clear zone of proteolytic activity, which corresponded to the band obtained with SDS-PAGE indicating that this could be a single-polypeptide enzyme.

Comparison of adsorbent with varying arm length and ligand density for the purification of recombinant hepatitis B virus surface antigen / 生物工程学报

Novel hydrophobic absorbents were synthesized by immobilizing butyl derivative onto the highly cross-linked agarose beads manufatured in China, which are used as matrix. The effect of the spacer arm length (3C, 8C and 10C) and ligand density (from 13 to 45 micromol/mL) on the hydrophobicity were investigated using purified Hepatitis B surface antigen (HBsAg) expressed by CHO cell lines. Also considering the effects of salt concentration and pH on HBsAg recovery and purification factor, orthogonal experiment design method was used to evaluated the absorbents. The results showed the butyl-S absorbent with the spacer arm length of C8, the ligand density of 22 micromol/mL gel showed the best performance for the separation of HBsAg. Approximately 100% HBsAg recovery and 60 as purification fold were achieved by this media under the operating condition of pH 7.0 and 9% of salt concentrateion.

Development of a new hydrophobic interaction chromatography absorbent and its application to the purification of recombinant hepatitis B surface antigen / 生物工程学报

A new hydrophobic absorbent based on homemade highly cross-linked agarose beads was synthesized by immobilizing butyl derivative onto the matrix linkage. The density of ligand was controlled by adjusting the concentration of butanethiol and the synthesis route was optimized by evaluating the purification efficiency of recombinant Hepatitis B surface antigen (HBsAg) expressed by Chinese hamster ovary (CHO) cell line. A high performance absorbent was finally screened out with up to 80% of HBsAg recovery and purification-fold (PF) about 20. Furthermore, the column pressure was about 0.06 MPa under the flow rate of 500cm/h, and no leaked butyl were detected after exposing the gel in common buffers, chaotropic agents, high concentrations of denaturing agents such as guanidine hydrochloride, urea and polar organic solvents. These results demonstrated that the absorbent have high physico-chemical stability, so it was available for the downstream process. Finally, after scaled up to 2L wet gel/batch, the absorbent was applied to the integration of three-step chromatography and obtained the purified CHO-HBsAg with 95% purity by SDS-PAGE and HPLC, which meet the requirements of SFDA. The purification efficiency and the reproducible ability of the absorbents were also evaluated from batch-to-batch. The results demonstrated that the absorbent met the requirement of scalable, reproducible, economic effect as well. This absorbent is a promising alternative exported HIC gel for wildly being used in Chinese pharmaceutical industries.

Rabbit IgG antibodies against Phospholipase A2 from Crotalus durissus terrificus neutralize the lethal activity of the venom

Crotalus durissus terrificus (C.d.t.) (South American rattlesnake) venom possesses myotoxic and neurotoxic activities, both of which are also expressed by crotoxin, the principal toxin of this venom. Crotoxin contains a basic phospholipase A2 (PLA2) and a non toxic acidic protein, crotapotin. We have produced and investigated the ability of IgG antibodies raised in rabbits against PLA2 to neutralize the lethality of the whole venom. PLA2 was isolated by gel filtration chromatography (Sephadex G-75). Specific antibodies were obtained by subcutaneous and intramuscular inoculation of PLA2 (700 µg) with Freund adjuvant. Groups of six mice (20 + 2 g) were inoculated with 0.5 ml i.p. of C. d. t. venom (4 µg) or a mixture of venom that had been preincubated with the desired volume of IgG antibodies. Mortality, recorded 24 and 48 h after inoculation, showed that IgG anti-PLA2 were more effective than anticrotalic serum in neutralizing the lethal activity. These results demonstrate that it could be possible to obtain an anti-venom made by specific antibodies with a high level of protection against the lethal component of C.d.t. venom, and/or the inclusion of these antibodies as a supplement in heterologous anti-venoms

El veneno de Crotalus durissus terrificus (C.d.t.) (Cascabel de Sud América) posee actividad miotóxica y neurotóxica, actividades que también exhibe el complejo crotoxina, principal componente tóxico de este veneno. El complejo crotoxina está constituido por una fosfolipasa A2 básica (PLA2) y una proteína acídica no tóxica, el crotapotín. En este trabajo se estudió la capacidad neutralizante de anticuerpos IgG anti-PLA2 sobre la letalidad inducida por el veneno entero. El antígeno PLA2, fue aislado por cromatografía de filtración en gel (Sephadex G-75). Se inocularon conejos machos por vía subcutánea e intramuscular, con 700 µg de PLA2 y adyuvante para la obtención de anticuerpos específicos. La capacidad neutralizante del antisuero se analizó en ratones por inoculación con diluciones de veneno entero preincubado con un volumen adecuado de anticuerpos IgG anti-PLA2. Se inocularon ratones controles con 0.5 ml i.p. de veneno (4 µg.ml-1). El número de muertes fue contabilizado a las 24 y 48 h posteriores a la inoculación, demostrándose que la capacidad neutralizante de los anticuerpos IgG anti-PLA2 fue superior a la obtenida con el antiveneno crotálico. Los resultados obtenidos demuestran la potencial aplicación de antivenenos constituidos por anticuerpos específicos contra PLA2, y/o la inclusión de estos anticuerpos como suplementos en antivenenos polivalentes

Studies on purification of methamidophos monoclonal antibodies and comparative immunoactivity of purified antibodies / 生物医学与环境科学（英文）

<p><b>OBJECTIVE</b>To purify Methamidophos (Met) monoclonal antibodies with two methods and compare immune activity of purified antibodies.</p><p><b>METHOD</b>Caprylic acid ammonium sulphate precipition (CAASP) method and Sepharose protein-A (SPA) affinity chromatography method were used to purify Met monoclonal antibodies, UV spectrum scanning was used to determine protein content and recovery of purified antibodies, sodium dodecylsulphate polyacrylamide gel electrophoresis (SDS-PAGE) was used to analyze the purity of purified antibodies, and enzyme-linked immunosorbent assay (ELISA) was used to determine immune activity of purified antibodies.</p><p><b>RESULTS</b>Antibody protein content and recovery rate with CAASP method were 7.62 mg/mL and 8.05% respectively, antibody protein content and recovery rate with SPA method were 6.45 mg/mL and 5.52% respectively. Purity of antibodies purified by SPA method was higher than that by CAASP method. The half-maximal inhibition concentration (IC50) of antibodies purified by SPA to Met was 181.26 microg/mL, and the linear working range and the limit of quantification (LOD) were 2.43-3896.01 microg/mL and 1.03 microg/mL, respectively. The IC50 of antibodies purified by CAASP to Met was 352.82 microg/mL, and the linear working range and LOD were 10.91-11412.29 microg/mL and 3.42 microg/mL, respectively.</p><p><b>CONCLUSION</b>Antibodies purified by SPA method are better than those by CAASP method, and Met monoclonal antibodies purified by SPA method can be used to prepare gold-labelled testing paper for analyzing Met residue in vegetable and drink water.</p>

Purification and properties of antiviral proteins from the leaves of Bougainvillea xbuttiana.

A non-phytotoxic, resistance inducing, proteinaceous antiviral principle was purified by ammonium sulphate fractionation, ion exchange chromatography and gel filtration from the leaves of Bougainvillea xbuttiana. It imparted resistance against tobacco mosaic virus (TMV) and sunnhemp rosette virus (SRV) in their respective test hosts viz. Nicotiana glutinosa, N. tabacum var. Samsun NN, and Cyamopsis tetragonoloba, respectively. The purified principle eluted as a single peak upon gel filtration, but exhibited two polypeptides on SDS-PAGE with Mr 28,000 and 24,000. The two polypeptides were found to be highly basic, rich in lysine with pI around 10.0 and 10.5, respectively. Since this principle effected local lesion inhibition in both treated and untreated top leaves of test host, it might be acting in the initial stages of virus infection as a systemic inducer.

Purification and characterization of Trypsin-Like Enzyme from the Pyloric Caeca of Cod(Gadus morhua) II

A tripsina do ceco pilórico do bacalhau (Gadus morhua) foi purificada por cromatografia deCHOM Sepharose 4B. Algumas características foram determinadas como atividade catalítica para T.A.M.E., substratos enzimáticos e inibidores de proteases. A enzima mostrou ponto isoelétrico de 5,30 e 5,89 e composiçäo de aminoácidos similar à tripsina bovina, mas diferiu por ter um alto percentual de aminoácidos ácidos e baixo valor em aminoácidos básicos. A tripsina do bacalhau também hidrolisou substratos de proteínas de pescados

Characterisation of Jack fruit lectin.

Jack fruit (Artocarpus Heterophyllus) seed extract contains a lectin termed Jack fruit lectin (JFL) which possesses diversed biological properties. A detailed analysis of its properties has been lacking. The present investigation was initiated to study the detail properties of JFL. After extraction and purification on affigel galactosamine-agarose column, JFL was subjected to ND-PAGE. Several different charged species from ND-PAGE upon SDS-PAGE gave rise to two dissimilar trimeric subunit at 12.5 and 15.0 KDa and retain biological activity. It was possible to elute the subunit bands separately from polyacrylamide gel to investigate their biological activity. Each subunit was found to be retained the lectin activity. Agglutinating activity of smaller subunit was found to be more, may be due to the greater amount of the subunit. This also suggests that each unit of trimeric JFL have similar lectin activity.

Production of Soluble VEGF Receptor Mutants for Inhibition of Angiogenesis / 대한암학회지

PURPOSE: Vascular endothelial growth factor (VEGF) is a potent angiogenic factor of many solid tumors, promoting vascularization and formation of metastases. In an attempt to generate effective VEGF inhibitors, the authors constructed the VEGF receptor mutants, expressed in E. coli and Sf9 insect cells, and examined their binding to VEGF. MATERIALS AND METHODS: The cDNA fragment encoding FLT-1 extracellular domain was cloned from human umbilical vein endothelial (HUVE) cell total RNA using RT-PCR. PCR- subcloning was performed using this template, in order to generate the deletion mutants by introducing FLT-1 partial sequences into E.coli expression vector pET-21d and baculovirus transfer vactors, pBAC-1 and pBAC-3. Two mutant proteins from baculovirus-infected insect cells were purified by heparin sepharose chromatography and immobilized into nitrdegrees Cellulose membrane followed by 125I-VEGF binding assay. RESULTS: Two mutant receptors, sFLT (1~7) and sFLT (2~4) expressed in E.coli appeared in inclusion body as insoluble proteins. The soluble mutant receptors were produced in low yield by baculovirus/insect cell expression system. Both immobilized mutant receptors, sFLT (1~7) and sFLT (2~4) were able to bind VEGF. CONCLUSION: These results suggest that a small soluble mutant receptor, sFLT (2~4), as well as sFLT (1~7) may be used effectively for bldegrees Cking angiogenic function of VEGF.

Cloning, expression and purification of recombinant streptokinase: partial characterization of the protein expressed in Escherichia coli

We cloned the streptokinase (STK) gene of Streptococcus equisimilis in an expression vector of Escherichia coli to overexpress the profibrinolytic protein under the control of a tac promoter. Almost all the recombinant STK was exported to the periplasmic space and recovered after gentle lysozyme digestion of induced cells. The periplasmatic fraction was chromatographed on DEAE Sepharose followed by chromatography on phenyl-agarose. Active proteins eluted between 4.5 and 0 percent ammonium sulfate, when a linear grandient was applied. Theree major STK derivatives of 47.5 kDa, 45 kDa and 32 kDa were detected by Western blot analysis with a polyclonal antibody. The 32-kDa protein formed a complex with human plasminogen but did not exhibit Glu-plasminogen activator activity, as revealed by a zymographic assay, whereas the 45-kDa protein showed a Km = 0.70 muM and kcat = 0.82 s(-1), when assayed with a chromogen-coupled subtrate. These results suggest that these proteins are putative fragments of STK, possibly derived from partial degradation during the export pathway or the purification steps. The 47.5-kDa band corresponded to the native STK, as revealed by peptide sequencing.

An alternative column chromatographic process for the production of human albumin

We have modified a standard column chromatography method for the preparation of albumin from human plasma. The proposed method utilizes Sephadex G-25, euglobulin precipitation, DEAE-Sepharose, ethanol heat treatment and Sephacryl S-200 HR. The procedure differs from that normally used by the introduction of Schneider's ethanol/heat treatment step and the elimination of a CM-Sepharose step, after the DEAE-Sepharose step. The proposed method was used to produce 15 batches of albumin, 100 liters of plasma per batch. Purity of more than 99 percent was obtained at an average yield of 26.5 g/l plasma for the albumin produced. All batches presented 99.2 to 99.9 percent monomer and 0.1 to 0.8 percent dimer, with no larger polymers detected. The utilization of the final gel filtration step eliminated highly polymerized albumin usually obtained during the process. The advantages of the proposed method are reduction in the overall process time from 6 to 5 days, elimination of the CM-Sepharose step and the reduction of 6 to 4 columns in series for the Sephacryl S-200 HR step.

Porphyrinogen carboxylyase. Studies on existence of isoenzymes

Porphyrinogen carboxylyase from normal rat liver was subjected to purification methods. Two different purification protocols were used. In both cases, the inital steps consisted in obtaining a liver homogenate, followed by centrifugation, salt precipitation and phosphate gel absorption. Scheme I consisted in then submiting the preparation to DEAE-cellulose, followed by Sephacryl S-200 and Phenyl-sepharose sequential column chromatographies. Scheme II involved an affinity column followed by a Sephadex G-75 gel filtration column. In both cases, the enzyme was stored at-20 degrees Celsius until its assay. The addition of 2mM dithiotreytol to the incubation media or to the enzyme extract before storage, did not help improve the activity nor the stability of the enzyme. Those fractions containing the maximal enzyme activity, detected using Uroporphyrinogen III or Pentacarboxy-porphyrinogen III as substrate, were not alawys present in the same tubes for the different columns employed. In addition, the degree of purification obtained in some steps was different according to the substrate employed. The results suggest the existence of at least two isoenzymes for rat liver porphyrinogen carboxy-lyase.

The rat thymus contains a heparin-binding factor that modulates steroidogenesis in the testis

Thymus development and function are under the influence of hormones secreted by the gonads and pituitary. On the other hand, thymurus is crucial for the development of reproductive capacitities in female and male rats and we have shown that a factor derived from the prepubertal rat thy mus has antigonadotropic effect in ovarian and testis cells in vitro. In the present paper we show that the rat thymic factor which modulates gonadotropin action in the gonads is an heparin-binding factor. This capacity was also used as a useful tool to obtain this activity from semipure extracts. An acetone extract was prepared from 15 day old male rats and subjected to molecular filtration chromatography. The activity, of those fractions was investigated in a testis cells biossay, by measuring testosterone secretion under basal and hCG-stimulation. Active fraction were processed in a heparin-Sepharose affinity column. We found that fractions that eluted with 0.6 and 2M NaCl/10mM Tris had biological specific activity. The electrophoretic procedure showed that the apparent molecular weight of the Heparin Sephadex binding factor is 60 kDa. Since this factor was obtained from a protein peak that eluted in the volume of carbonic anhidrase a dimerization process could be involved. Present results show that the rat thymus has an heparin-binding factor that interacts with hCG in testis cells. This factor could play an interesting role in the mutual influence between thymus and gonads.

Distribution of phospholipase C isozymes in normal human lung tissue and their immunohistochemical localization

Phospholipase C(PLC) plays a central role in signal transduction and it is important in cellular growth, differentiation and transformation. There are currently ten known mammalian isozymes of PLC identified and cloned. However, there are no report of PLC distribution in human lung tissue or their significances in pulmonary diseases. Presence of various PLC isozymes in normal human lung tissue was studied from surgical specimens. PLC isozymes in tissue extracts of the lung were partially purified by successive chromatographic steps on heparin-sepharose CL-6B conventional and TSKgel heparin-5PW HPLC columns and their activities were assayed. PLC activity peaks identified in the chromatography were immunoblotted with specific antibodies against ten known mammalian PLC isozymes(PLC-beta 1-4, -gamma 1-2, and -delta 1-4). In addition, immunohistochemical staining of the lung tissue was performed to determine subcellular and histological localization of PLC isozymes. The results indicate that normal human lungs contain beta 1, beta 3, gamma 1, and delta 1, isozymes of PLC. The order of amount present in the lung tissue was PLC-delta 1 > gamma 1 >beta 1 >> beta 3, in descending order. On immunohistochemistry, PLC-gamma 1 was most widely distributed and was present in bronchiolar epithelium, in type I and type II pneumocytes as well as in fibroblasts of the interstitial tissue. PLC-delta 1 was present in the cytoplasm of the bronchiolar epithelium whereas PLC-beta 1 was localized to the apical membranous portion of the same epithelium. PLC-beta 3 was seen in the nucleus of the respiratory and alveolar lining epithelium as well as in the nucleus of lung fibroblasts.