Fish Species Identification by Isoelectric Focusing: Sarcoplasmic Protein Polymorphism in Monkfish (Lophius americanus)

Abstract Monkfish (Lophius americanus) sarcoplasmic protein patterns were found to be polymorphic with respect to separations using isoelectric focusing. Reproducible protein pattern variations were not detected using cellulose acetate or polyacrylamide gel disc electrophoresis. Monkfish sarcoplasmic proteins separated on pH 3.5-9.5 Ampholine PAGplates or on pH 2.5-9.0 agarose IEF gels yielded similar patterns showing 3 distinct, reproducible variations. On close examination, the pH 3.5-9.5 Ampholine PAGplate patterns could be further subdivided into 3 additional variations. A high resolution pH 3.5-5.0 agarose IEF gel was able to resolve a total of 10 different monkfish pattern variations in a sample of 24 individuals. A model was proposed suggesting the existence of 16 distinct variations in the monkfish sarcoplasmic protein pattern, based on the various combinations of 4 protein bands. In identifying samples of monkfish meat, it is necessary to compare the unknown pattern with the possible variant patterns to effect a reliable identification. On recommendation by the Associate Referee, the method for fish species identification based on polyacrylamide gel isoelectric focusing, 18.A01-18.A04, has been adopted official final action with no restrictions as to the species that may be identified.

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Rapid Fish Species Identification by Agarose Gel Isoelectric Focusing of Sarcoplasmic Proteins

Journal of AOAC INTERNATIONAL ◽

10.1093/jaoac/64.1.38 ◽

1981 ◽

Vol 64 (1) ◽

pp. 38-43

Author(s):

Ronald C Lundstrom

Keyword(s):

Species Identification ◽

Isoelectric Focusing ◽

Fish Species ◽

Protein Pattern ◽

Agarose Gel ◽

Tissue Fluid ◽

Protein Patterns ◽

Fish Species Identification ◽

Species Specific ◽

Gel Isoelectric Focusing

Abstract A rapid method is described for fish species identification by agarose gel isoelectric focusing (AGIEF). The AGIEF method can be completed in less than 2 h and gives reproducible species-specific sarcoplasmic protein patterns. Protein patterns are similar using either centrifuged tissue fluid or muscle tissue as the sample. One species, monkfish (Lophius americanus), has a polymorphic protein pattern. A predominant pattern was found in 66.7% of the individuals; 2 variant patterns were equally distributed among the remaining 33.3%. AGIEF offers a more rapid, less expensive alternative to the current AOAC official first action method for fish species identification based on polyacrylamide gel isoelectric focusing.

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Fish Species Identification by Agarose Gel Isoelectric Focusing: Collaborative Study

Journal of AOAC INTERNATIONAL ◽

10.1093/jaoac/66.1.123 ◽

1983 ◽

Vol 66 (1) ◽

pp. 123-127

Author(s):

Ronald C Lundstrom ◽

◽

M C Annand ◽

C R Barmore ◽

S A Braddon ◽

...

Keyword(s):

Species Identification ◽

Isoelectric Focusing ◽

Fish Species ◽

Protein Pattern ◽

Collaborative Study ◽

Agarose Gel ◽

Protein Patterns ◽

Unknown Protein ◽

Fish Species Identification ◽

Gel Isoelectric Focusing

Abstract A method for fish species identification by agarose gel isoelectric focusing was collaboratively studied to determine if unknown samples could be identified by comparison of the unknown protein pattern with a photograph of protein patterns from authentic species. Fourteen collaborators were sent 19 or 20 unknown samples along with supplies and instructions for making pH 2.5-9.0 agarose isoelectric focusing gels. The unknown sarcoplasmic protein patterns were identified by comparison with a supplied 8 X 10 in. photograph of protein patterns from 18 authentic species. The 14 collaborators identified a total of 276 unknown samples with 84% accuracy. Eight of 14 collaborators averaged 90% or more correct. Two collaborators averaged 80-85% correct, while the remaining 4 collaborators averaged 50-69% correct. No one species was consistently misidentified; however, the method is not recommended for adoption in its present form.

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Fish Species Identification By Thin Layer Polyacrylamide Gel Isoelectric Focusing: Collaborative Study

Journal of AOAC INTERNATIONAL ◽

10.1093/jaoac/63.1.69 ◽

1980 ◽

Vol 63 (1) ◽

pp. 69-73 ◽

Cited By ~ 1

Author(s):

Ronald C Lundstrom ◽

◽

C R Barmore ◽

J Krzynowek ◽

A F LaTerza ◽

...

Keyword(s):

Species Identification ◽

Isoelectric Focusing ◽

Fish Species ◽

Protein Pattern ◽

Collaborative Study ◽

Protein Patterns ◽

Unknown Species ◽

Fish Species Identification ◽

Gel Isoelectric Focusing ◽

Standard Protein

Abstract A method for fish species identification by isoelectric focusing of sarcoplasmic proteins on LKB Ampholine PAGplates, pH 3.5–9.5, was collaboratively studied to determine whether photographs of protein patterns from authentic species could be used to identify unknown samples. Seven collaborators were sent 8 unknown samples in duplicate to identify from an 8 × 10 in. photograph of an Ampholine PAGplate showing standard protein patterns from 14 species of fish. The 7 collaborators identified the unknown samples with 93% accuracy. Eight of 14 monkfish samples were not identified correctly and showed protein patterns different from the protein patterns shown in the photographic standard. All of the incorrectly identified samples showed the same protein pattern. The other 7 unknown species were all correctly identified. The method has been adopted as official first action.

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Fish Species Identification by Thin Layer Agarose Isoelectric Focusing and Densitometric Scanning

Journal of AOAC INTERNATIONAL ◽

10.1093/jaoac/65.1.119 ◽

1982 ◽

Vol 65 (1) ◽

pp. 119-122

Author(s):

Warren D Hamilton

Keyword(s):

Thin Layer ◽

Species Identification ◽

Isoelectric Focusing ◽

Fish Species ◽

Visible Range ◽

Protein Patterns ◽

Reliable Technique ◽

Support Medium ◽

Fish Species Identification ◽

Gel Isoelectric Focusing

Abstract Examination of sarcoplasmic proteins of fish by thin layer polyacrylamide gel isoelectric focusing as a means of fish species identification is a powerful and reliable technique, but it displays a number of disadvantages. These problems include the care required when handling acrylamide monomer (a neurotoxin), the mechanical skill needed in molding the gel, the difficulties in ensuring correct gel polymerization, and the extensive destaining periods. Specially treated agarose has been used to obtain protein patterns for an interval of pH 5-8. The patterns so produced were scanned by a densitometer in the visible range after completing staining and destaining. Most of the problems associated with polyacrylamide gels have been overcome by using agarose is a support medium.

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Fish Species Identification by Thin Layer Isoelectric Focusing

Journal of AOAC INTERNATIONAL ◽

10.1093/jaoac/62.3.624 ◽

1979 ◽

Vol 62 (3) ◽

pp. 624-629

Author(s):

Ronald C Lundstrom

Keyword(s):

High Resolution ◽

Thin Layer ◽

Isoelectric Focusing ◽

Fish Species ◽

Protein Separation ◽

Protein Pattern ◽

Polyacrylamide Gels ◽

Protein Patterns ◽

Sample Application ◽

Gel Isoelectric Focusing

Abstract Conventional electrophoretic techniques generally lack the resolution and reproducibility needed for the reliable identification of fish species. Variations in stabilizing media composition, sample application technique, separation time, applied voltage or current, and the analyst’s skill all affect the protein pattern. Thin layer polyacrylamide gel isoelectric focusing (TLIEF), a high resolution protein separation technique, has been applied to the identification of fish species. Sarcoplasmic proteins are separated according to their isoelectric points in a stable, reproducible ptT^gradient. Protein patterns for 12 species of fish are compared in 4.0% polyacrylamide gels with pH 4.0–6.0 and pH 3.5–10 gradients. Similar patterns are shown in commercially prepared 5.0% polyacrylamide gels with pH 4.0–6.5 and pH 3.5–9.5 gradients (LKB PAG plates). The protein patterns are reproducible in each pH gradient and also correlate well between user-prepared and commercially prepared gels. The inherent high resolution and excellent reproducibility of TLIEF should allow the positive identification of fish species without the costly procedure of using known species as standards.

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Collaborative Study of a Rapid Electrophoretic Method for Fish Species Identification

Journal of AOAC INTERNATIONAL ◽

10.1093/jaoac/52.4.703 ◽

1969 ◽

Vol 52 (4) ◽

pp. 703-707

Author(s):

Robert J Learson

Keyword(s):

Cellulose Acetate ◽

Species Identification ◽

Fish Species ◽

Collaborative Study ◽

Correct Identification ◽

Protein Patterns ◽

Electrophoretic Method ◽

Fish Species Identification ◽

Fish Samples ◽

Standard Protein

Abstract A rapid electrophoretic method for fish species identification, using cellulose acetate as the supporting medium, was collaboratively studied to determine whether photographs of standard protein patterns from authentic species could he used to identify unknown samples. Twelve collaborators were sent six unknown samples of fish in duplicate to identify from a set of photographs representing standard patterns from nine species of fish. Results obtained from the 10 reporting collaborators indicated that correct identification from photographic standards was extremely difficult. Although the analysts were able to match the duplicates with an accuracy of 90%, only 39% of the unknowns were correctly identified. It is recommended that the method be collaboratively studied using authentic fish samples for standards instead of photographic standards.

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