Preparation of Samples for Chemical and Biochemical
PARTNERSHIP FOR THE DELAWARE ESTUARY Science Group
Preparation of Samples for Chemical and Biochemical Characterization of Algae and Seston Date Prepared: 2/16/1994
Prepared By: __ Danielle Kreeger______
Suggested Citation: Kreeger, D. 1994. Preparation of Samples for Chemical and Biochemical Characterization of Algae and Seston. Partnership for the Delaware Estuary. PDE Method No. 04. 3pp. PDE-Method (2/16/1994)
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Preparation of Samples for Chemical and Biochemical Characterization of Algae and Seston Partnership for the Delaware Estuary (PDE) Method Danielle Kreeger Description This method organizes the various analyses of the chemistry and biochemistry of either microalgae or seston samples. Detailed methods for each analysis are described in other documents. Use this procedure for algae cultured in glassware and using culture methods (request DK-SOP-01 and DK-SOP-02) Procedures 4.1. Harvest 2 l of algae (or the appropriate amount for analysis) from culture vessel (request DK-SOP-02) or collect at least 2 L of water from a field site. Keep samples cool but do not freeze and within 12 h (preferably within 1-2 h), mix and divide sample as follows. 4.2. Add exactly 10 ml algae to a 20 ml LSC vial (x15) 4.2.1 Use 3 repls. to enumerate cell concentration using either a hemacytometer or Coulter Counter. If counting is delayed, fix with acid Lugol's solution. 4.2.2 Sonicate 12 samples for 20-30 min and freeze at -70oC .until analysis. Use 4 replicates to perform protein analysis (PDE-Method-12 Biochemical Analyses I. Protein). Store the remaining 8 replicates for other potential analyses (amino acids, fatty acids). 4.3 Filter a known volume (e.g. 25-100 ml) of algae/seston through a prepared GF/F filter (see above for preparation); x19 plus blanks. Blanks are formed by centrifuging an additional 12 vols. of each sample at 3000 rpm for 15 min, and collecting the supernatants (=blanks). 4.3.1. Transfer 15 ml of 4 filtrates and 15 ml of 4 blanks to polystyrene test tubes and freeze at -70oC. Use 4 sample and 4 blank filters and blank filtrates for phosphorus analyses if desired. 4.3.2. Freeze-dry filters in disposable polystyrene test tubes. Perform the following different analyses: 4.3.2.1 Use 4 filters and 4 blanks for analysis of dry and ash-free dry weight (PDEMethod-07 Biomass Measurement II. Dry and Ash-Free Dry Weight). 4.3.2.2 Use 4 filters and 4 blanks for analysis of lipids (PDE-Method-13 Biochemical Analyses II. Lipid).
PDE-Method (2/16/1994)
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4.3.2.3 Use 4 filters and 4 blanks for analysis of carbohydrate (PDE-Method-14 Biochemical Analyses III. Carbohydrates). 4.3.2.2 Use 4 filters for analysis of chlorophyll if desired. 4.4. Freeze 90 ml of each centrifuged blank (x3) for nitrogen analysis. 4.5. Filter 15 ml of each sample (x3) and 15 ml of each centrifuged blank (x3) through a 0.45 μm polycarbonate. Freeze filters and filtrates in separate polystyrene tubes at -70oC for silicon analysis if desired.
PDE-Method (2/16/1994)
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Preparation of Samples for Chemical and Biochemical Characterization of Algae and Seston Date Prepared: 2/16/1994
Prepared By: __ Danielle Kreeger______
Suggested Citation: Kreeger, D. 1994. Preparation of Samples for Chemical and Biochemical Characterization of Algae and Seston. Partnership for the Delaware Estuary. PDE Method No. 04. 3pp. PDE-Method (2/16/1994)
Page 1
Preparation of Samples for Chemical and Biochemical Characterization of Algae and Seston Partnership for the Delaware Estuary (PDE) Method Danielle Kreeger Description This method organizes the various analyses of the chemistry and biochemistry of either microalgae or seston samples. Detailed methods for each analysis are described in other documents. Use this procedure for algae cultured in glassware and using culture methods (request DK-SOP-01 and DK-SOP-02) Procedures 4.1. Harvest 2 l of algae (or the appropriate amount for analysis) from culture vessel (request DK-SOP-02) or collect at least 2 L of water from a field site. Keep samples cool but do not freeze and within 12 h (preferably within 1-2 h), mix and divide sample as follows. 4.2. Add exactly 10 ml algae to a 20 ml LSC vial (x15) 4.2.1 Use 3 repls. to enumerate cell concentration using either a hemacytometer or Coulter Counter. If counting is delayed, fix with acid Lugol's solution. 4.2.2 Sonicate 12 samples for 20-30 min and freeze at -70oC .until analysis. Use 4 replicates to perform protein analysis (PDE-Method-12 Biochemical Analyses I. Protein). Store the remaining 8 replicates for other potential analyses (amino acids, fatty acids). 4.3 Filter a known volume (e.g. 25-100 ml) of algae/seston through a prepared GF/F filter (see above for preparation); x19 plus blanks. Blanks are formed by centrifuging an additional 12 vols. of each sample at 3000 rpm for 15 min, and collecting the supernatants (=blanks). 4.3.1. Transfer 15 ml of 4 filtrates and 15 ml of 4 blanks to polystyrene test tubes and freeze at -70oC. Use 4 sample and 4 blank filters and blank filtrates for phosphorus analyses if desired. 4.3.2. Freeze-dry filters in disposable polystyrene test tubes. Perform the following different analyses: 4.3.2.1 Use 4 filters and 4 blanks for analysis of dry and ash-free dry weight (PDEMethod-07 Biomass Measurement II. Dry and Ash-Free Dry Weight). 4.3.2.2 Use 4 filters and 4 blanks for analysis of lipids (PDE-Method-13 Biochemical Analyses II. Lipid).
PDE-Method (2/16/1994)
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4.3.2.3 Use 4 filters and 4 blanks for analysis of carbohydrate (PDE-Method-14 Biochemical Analyses III. Carbohydrates). 4.3.2.2 Use 4 filters for analysis of chlorophyll if desired. 4.4. Freeze 90 ml of each centrifuged blank (x3) for nitrogen analysis. 4.5. Filter 15 ml of each sample (x3) and 15 ml of each centrifuged blank (x3) through a 0.45 μm polycarbonate. Freeze filters and filtrates in separate polystyrene tubes at -70oC for silicon analysis if desired.
PDE-Method (2/16/1994)
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