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Systematic screening and cloning of phosphoproteins from human cardiac sarcomeres by in-gel digestion and mass spectrometry. The molecular machinery by which sarcomeres regulate contraction and cardiac pump function is not fully understood and, consequently, it is difficult to target these processes therapeutically. Phosphorylation of sarcomeric proteins may play a role in this regulation and has been explored in some detail using Western blots with synthetic peptides that correspond to known sequences of phosphorylated proteins. We therefore undertook to use proteomic approaches to gain a more complete picture of phosphoproteins involved in cardiac function. Cardiac myofibrils were isolated from human left ventricle using a modified method of cleavage and shearing, and subjected to an isolation procedure which retains intact sarcomere integrity. Phosphoproteins were purified from these myofibrillar preparations by an immunoaffinity step and then were subsequently subjected to sequencing by microcapillary liquid chromatography-electrospray ionization-tandem mass spectrometry (microLC-ESI-MS-MS). Data were obtained from three technical replicates of each of three separate purifications and these data were searched against the Swiss-Prot database. More than 100 unique phosphoproteins were identified, with 54 successfully identified by fragmentation spectra. Phosphorylation sites were detected in 39 of these proteins, and where possible, phosphorylated residues were localized to cysteine residues by comparing the precursor and fragment ion series of the phosphopeptides. A phosphoprotein-specific enzymatic hydrolysis of the biological samples by protein phosphatase 2A indicated that 17 were actually regulated by phosphorylation. Four of these were cardiac troponin I (TnI), myosin regulatory light chain, and multiple titin isoforms. Phosphorylation of titin or myosin regulatory light chain could indicate a role for this type of regulation in cardiac pump function. Identification of many phosphoproteins with unique sites should facilitate direct targeting of particular signaling pathways.Highly sensitive human papilloma virus detection by PCR combined with time-resolved fluoroimmunoassay. Detection of high-risk subtypes of human papilloma virus (HPV), particularly those involved in the pathogenesis of cervical cancer, is the major goal of HPV diagnosis. The present paper describes time-resolved fluoroimmunoassay (TRFIA) as
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If this is your first visit, be sure to check out the FAQ by clicking the link above. You may have to register before you can post: click the register link above to proceed. To start viewing messages, select the forum that you want to visit from the selection below. In this documentation, «minimal API» refers to the fact that the actual running state of the contract is on the blockchain, and all required data is in the blockchain. No data is stored on the Ethereum blockchain. Thus, there is no need to retrieve accounts or state manually from the blockchain. Parity’s minimal API allows any existing Ethereum library to call Parity as it is defined in their API. GetInitialTransaction This is the method used for obtaining the transaction hash that is used to initialize the contract account’s data. Properties: contract_address : Address, the address of the contract source_account : Address, an account from which the transaction was initialized gas_limit : Number, the available gas for the transaction gas_price : Number, a gas price to use, in wei, for the transaction value : Number, the value of the transaction to execute, in wei fees : Amount, the gas price multiplied by the total gas of the transaction, in wei transaction_id : String, a unique identifier for the transaction Returns: an object with the transaction’s information, e.g. in the following example: Note: If gas_limit is zero, the transaction will be orphaned and dropped. In the event that gas_limit is less than gas_price and a transaction with the same contract_address has not been already included in a block, Parity will use a default gas limit and charge the gas price per block for the transaction. Examples getInitialTransaction() setter Initiate a state transition of the current contract account. This call is equivalent to executing a transaction with signature, source_account, gas_limit, and gas_price. setInitialTransaction_source_account(account) setInitialTransaction_gas_limit(gas_limit) setInitialTransaction_gas_price(gas_price) setInitialTransaction_value(value) setInitialTransaction_fees(fees) setInitialTransaction_transaction
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